Plasmodium falciparum biology
Encyclopedia
Plasmodium falciparum
has been the focus of much research due to it being the causative agent of malaria
. This article describes some of the recent findings surrounding the unique biology of this organism.
has a complicated life-cycle, requiring both a human and a mosquito
host, and differentiating multiple times during its transmission/infection process.
is transmitted to humans by the females of the Anopheles
species of mosquito
. There are about 460 species of Anopheles
mosquito
, but only 68 transmit malaria. Anopheles gambiae
, found in Africa, is one of the best malaria vectors. It is long-lived, prefers feeding on humans, and lives in areas near human habitation.
Prior to transmission, Plasmodium falciparum
resides within the salivary gland of the mosquito
. The parasite is in its sporozoite stage at this point. As the mosquito
takes its blood meal, it injects a small amount of saliva into the skin wound. The saliva contains antihemostatic and anti-inflammatory enzymes that disrupt the clotting process and inhibit the pain reaction. Typically, each infected bite contains 5-200 sporozoites which proceed to infect the human vector. Once in the human bloodstream, the sporozoites only circulate for a matter of minutes before infecting liver cells.
sporozoites enter hepatocytes. At this point, the parasite loses its apical complex and surface coat, and transforms into a trophozoite. Within the parasitophorous vacuole of the hepatocyte
, P. falciparum
undergoes schizogonic development. In this stage, nucleus divides multiple times with a concomitant increase in cell size, but without cell segmentation. This exoerythrocytic schizogony stage of P. falciparum
has a minimum duration of roughly 5.5 days. After segmentation, the parasite cells are differentiated into merozoites.
After maturation, the merozoites are released from the hepatocytes and enter the erythrocytic portion of their life-cycle. Note that these cells do not reinfect hepatocytes.
Plasmodium possess only a single pyruvate dehydrogenase
enzyme (PDH) complex. This is localized to the plastid-like organelle known as the apicoplast
. Unlike most eukaryotes, Plasmodium lacks a mitochondrial PDH. The PDH catalyzes the conversion of pyruvate to acetyl-CoA
, an important precursor for the tricarboxylic acid cycle and type II fatty acid
synthesis.
The process of maturation within the liver is still being investigated. In the species Plasmodium yoelli the exit from the liver appears to be dependent on type II fatty acid synthesis. Deletions in either the pyruvate dehydrogenase E1 alpha and E3 subunits produce a phenotype similar to that found in mutants of the type II fatty acid synthesis pathway. These mutants appear normal in blood stage development, mosquito stage development and early liver stage development but fail to exit the liver cells.
Plasmodium is unable to synthesize sterol
s they must obtain these from the host. However manipulation of cholestoerol metabolism does not impede the development of the merozoites.
Invasion of the liver cell chances the properties of the cell itself. The cell membrane becomes rougher and the cell itself becomes significantly stiffer. The mechanisms of these changes is currently unknown.
Merozoite
After release from the hepatocytes, the merozoites enter the bloodstream prior to infecting red blood cells. At this point, the merozoites are roughly 1.5 μm in length and 1 μm in diameter, and use the apicomplexan invasion organelles (apical complex, pellicle and surface coat) to recognize and enter the host erythrocyte.
Unlike species in the genus Toxoplasma which have multiple rhopteries
, Plasmodium species typically only have two. These may be referred in the literature to as the 'paired organ'.
The mechanism of cell attachment and invasion are under active investigation. Several molecules have been implicated in this process. Two of these are the genes PfRh2a and PfRh2b. These are large proteins of about 3200 amino acids in length. They differ only in the last 500 amino acids of the C terminal and have clearly arisen by a process of gene duplication and mutation. The 500 amino acid region includes an ectodomain, a transmembrane domain and a cytoplasmic domain. PfRh2b is essential for a well-defined invasion pathway while PfRh2a is not required or sufficient for this pathway. It has been shown that the reason for this difference lies in the cytoplasmic domain.
The parasite first binds to the erythrocyte in a random orientation. It then reorients such that the apical complex is in proximity to the erythrocyte membrane. A tight junction is formed between the parasite and erythrocyte. As it enters the red blood cell, the parasite forms a parasitophorous vesicle, to allow for its development inside the erythrocyte.
Five separate invasion paths have been delineated. The most common pathway is neuraminidase
resistant, trypsin
sensitive and chymotrypsin
resistant invasion. Some parasites have a neuraminidase- and trypsin-sensitive phenotype indicating a dependence on the erythrocyte binding antigen 175/glycophorin A
pathway(s). Most isolates appear to be dependent on a trypsin sensitive pathway.
The mechanics of the binding process have been partly elucidated. The low potassium levels in the blood (3.5-5.0 millimoles per liter) activates a phospholipase C enzyme. This enzyme initiates a pathway that leads to a rise in the intracellular calcium. This rise in calcium triggers secretion of microneme proteins including the 175 kD erythrocyte binding antigen and apical membrane antigen 1
to the merozoite surface where they can bind to erythrocyte surface proteins including glycophorin A
.
It appears that the merozoite surface protein 1 (MSP1) binds to heparin
like molecules on the surface of the erythrocyte and ththat is binding is an essential step in the invasion process.
The protein PfRON2 via the C-terminal as well as the central cysteine-rich domain interacts with PfAMA1. This protein also appears to be involved in erythrocyte invasion.
Trophozoite
After invading the erythrocyte, the parasite loses its specific invasion organelles (apical complex and surface coat) and de-differentiates into a round trophozoite located within a parasitophorous vacuole in the red blood cell cytoplasm. The young trophozoite (or "ring" stage, because of its morphology on stained blood films) grows substantially before undergoing schizogonic division.
During asexual development the parasite increases in size to ~50% of the uninfected erythrocyte volume: the infected erythrocyte volume remains relatively constant. Haemoglobin content gradually decreases but its concentration remains constant until the early trophozoite stage when it decreases by 25%. It then remains constant again until just prior to rupture. During early sexual development the gametocyte has a similar morphology to a trophozoite but subsequently undergoes a dramatic shape change.
Within the red blood cell, the parasite metabolism depends greatly on the digestion of hemoglobin
. A set of enzymes known as plasmepsin
s which are aspartic acid proteases
are used to degrade hemoglobin
. The parasite digests 70-80% of the erythrocyte's haemoglobin but utilizes only ~15% in de novo protein synthesis. Intraerythrocytic Plasmodium falciparum utilizes only a fraction of the amino acids derived from the digestion of host cell cytosol for the biosynthesis of its proteins. The excess amino acids are exported from the infected erythrocyte by new transport pathways created by the parasite. The reason proposed for this apparently excessive digestion of haemoglobin is the colloid-osmotic hypothesis which suggests that the digestion of haemoglobin increases the osmotic pressure within the infected erythrocyte leading to its premature rupture and subsequent death of the parasite. To avoid this fate much of the haemoglobin is digested and exported from the erythrocyte. This hypothesis has been experimentally confirmed.
Infected erythrocytes are often sequestered in various human tissues or organs, such as the heart, liver and brain. This is caused by parasite derived cell surface proteins being present on the red blood cell membrane and it is these proteins that bind to receptors on human cells. Sequestration in the brain causes cerebral malaria, a very severe form of the disease, which increases the victim's likelihood of death.
The parasite can also alter the morphology of the red blood cell causing knobs on the erythrocyte membrane.
Erythrocyte invasion and growth leads to activation of several distinct anion channels and a non-selective Ca2+-permeable cation channel. The non-selective cation channel's activation allows entry of Ca2+ and Na+. Absence of the channels is incompatible with pathogen survial. Although the mechanism of activation of these channels is not know it is presumed to be due to oxidation stree generated by the parasite because similar or identical channels are activated by oxidation of non-infected erythrocytes. Ca2+ entry stimulates an intraerythrocytic scramblase
that facilitates bi-directional phospholipid
migration across the bilayer. This results in an alternation of the cell membrane's phosphatidylserine
asymmetry. Exposure of phosphatidylserine at the outer surface of the cell membrane is followed by binding to phosphatidylserine receptors on macrophage
s and the subsequent phagocytosis of the affected erythrocyte. It appears that the parasite because of its growth requirements is in a race to complete its life cycle before the infected erythrocyte is phagocytosed.
Schizont
At the schizont stage, the parasite replicates its DNA multiple times without cellular segmentation. These schizonts then undergo cellular segmentation and differentiation to form roughly 16-18 merozoite cells in the erythrocyte. The merozoites burst from the red blood cell, and proceed to infect other erythrocytes. The parasite is in the bloodstream for roughly 60 seconds before it has entered another erythrocyte.
This infection cycle occurs in a highly synchronous fashion, with roughly all of the parasites throughout the blood in the same stage of development. This precise clocking mechanism has been shown to be dependent on the human host's own circadian rhythm
. Specifically, human body temperature changes as a result of the circadian rhythm, seem to play a role in the development of P. falciparum
within the erythrocytic stage.
The synchronicity of the erythrocytic cycle is at least in part dependent on melatonin
secretion by the host. A mechanism for this has been proposed. Melatonin can activate phospholipase C which acts to generate inositol trisphosphate (IP3) which opens IP3 sensitive calcium
channels in the endoplasmic reticulum
. The released calcium in its turn controls the cycle through mechanisms that have yet to be understood.
host.
is taken up by the female Anopheles
mosquito
as it takes its bloodmeal from an infected human.
, the gametocytes leave the erythrocyte shell and differentiate into gametes. The female gamete maturation process entails slight morphological changes, as it becomes enlarged and spherical. On the other hand, the male gamete maturation involves significant morphological development. The male gamete's DNA
divides three times to form eight nuclei. Concurrently, eight flagella are formed. Each flagella pairs with a nucleus to form a microgamete, which separates from the parasite cell. This process is referred to as exflagellation.
Gametogenesis has been shown to be caused by: 1) a sudden drop in temperature upon leaving the human host, 2) a rise in pH within the mosquito
, and 3) xanthurenic acid
within the mosquito.
. The zygote
then develops into an ookinete. The zygote
and ookinete are the only diploid stages of P. falciparum
.
within the mosquito
. It traverses the peritrophic membrane of the mosquito
midgut and cross the midgut epithelium. Once through the epithelium, the ookinete enters the basal lamina, and forms an oocyst.
The processes of maturation and invasion of the mosquito gut are under investigation. Both chitinase
and plasmepsin
4 (an aspartic acid protease
) are known to be involved in the invasion process.
During the ookinete stage, genetic recombination can occur. This takes place if the ookinete was formed from male and female gametes derived from different populations. This can occur if the human host contained multiple populations of the parasite, or if the mosquito
fed from multiple infected individuals within a short time-frame.
A G-actin binding protein has been implicated in this process. These proteins - probably acting as dimers - bind actin monomers just before sporogony.
Azithromycin
has been shown to suppress apicoplast replication at the period of sporozoite production in oocysts.
The proteins enolase
and actin
are present on the surface of ookinetes but their function there, if any, is unknown.
The myosin gene (MyoA) class XIV is essential for ookinete motility in the mosquito. Mutants placed under a different promotor active in the blood stages failed to complete their life cycle. Disruption serine repeat antigen 5 blocks parasite inhibits egress of sporozoites from an oocyst.
bite.
s of Plasmodium sporozoites are linked to the parasite pellicle
via long tethering proteins. The tethers originate from the inner membrane complex and are arranged in a periodic fashion following a 32 nanometer repeat. The tethers pass through a subpellicular structure that encompasses the entire parasite, probably as a network of membrane associated filaments.
DNA synthesis begins in the relatively small trophozoites but nuclear subdivision, which leads to the formation of multinucleate cells, occurs only during schizogony. Whether or not any gap phases exist between each round of DNA synthesis and mitosis is unknown. Eventually, a schizont composed of 8–32 nuclei undergoes segmentation, which culminates with the formation of individual merozoites that burst from the erythrocyte into the blood stream.
sulfate and chondroitin
sulfate type membrane receptors on host cells.
Latency of sporozoites is controlled by the eIF2
alpha kinase IK2, a general inhibitor of protein synthesis. Puf2 participates in the regulation of IK2 and inhibits premature sporozoite transformation. In contrast Puf1 appears to be dispensable.
The RNA binding protein family PUF member Pumilio-2 (Puf2) appears to be involved in transformation of sporozoites into the hepatic stage of the life cycle. Knock out mutants of this gene exhibit genome wide transcriptional changes resulting in loss of gliding motility, cell traversal ability, reduction in infectivity and trigger metamorphosis typical of early Plasmodium intra-hepatic development.
The division of the liver stages into thousands of merozoites is a complex process. In parallel with nuclear division, the apicoplast and mitochondrion become two extensively branched and intertwining structures. The organelles subsequently undergo morphological and positional changes prior to cell division. Finally to form merozoites, the parasite undergoes cytokinesis.
Type II fatty acid
biosynthesis is vital for this stage in the life cycle. This pathway may be inhibited by the antibiotic triclosan
.
During this stage of development the sporozoite selectively discards organelles unnecessary for growth at this stage of the life cycle. Among these are the micronemes and the inner membrane complex.
The host iron regulatory hormone hepcidin
which is synthesised in the liver and spleen
, appears to be able to inhibit growth of the liver stages. Levels of this hormone are elevated during infection and seem to correlate with the anaemia often found in malaria. Erythrocytic parasitaemia, above a minimum threshold, impairs the growth of subsequent liver cell sporozoite infection. The production of hepcidin leads to the redistributes iron away from hepatocytes thus slowing the development of the iron dependent liver stage.
Liver hepcidin expression is upregulated and downregulated during the early and late stages of malaria infection respectively. Inflammation and erythropoietin, rather than the iron sensing pathway, are involved in the regulation of hepcidin expression. Treatment of malaria infected mice with anti hepcidin neutralizing antibodies increased parasitemia and mortality rates. Overexpression of hepcidin improves the outcome.
Two families of proteins are known to be involved in this process: the reticulocyte binding-like homologues (PfRh or PfRBP) and erythrocyte binding-like (EBL) proteins. The PfRh family consists of five proteins and a pseudogene
: PfRh1, PfRh2a, PfRh2b, PfRh3, PfRh4 and PfRh5. PfRh3 is a transcribed pseudogene in all strains examined to date. All the other members of this family bind to erythrocyes and antibodies to them inhibit invasion. PfRh5 is located within the rhoptries and appears to be an essential gene.
The EBL family of proteins includes EBA-175, EBA-181 (also known as JESEBL), EBA-140 (also known as BAEBL) and EBL-1. Whilst these parasite ligands function in merozoite invasion by binding to specific receptors on the erythrocyte, they appear also to have a central role in activation of the invasion process. Binding of EBA-175 to its receptor, glycophorin A
, restores the basal cytosolic calcium levels after interaction of the merozoite with the erythrocyte and triggers the release of rhoptry proteins.
There seems to be some overlap between the functions of these proteins. Loss of EBA-175 can be compensated by increased expression of PfRh4.
Some details of the invasion process are known. The rhoptery protein RON2 is inserted into the erythrocyte membrane. The protein AMA1 secreted from microneme then binds to RON2. RON2 forms part of a macromolecular complex which includes RON2, RON 4, RON5 and RON8.
The residues of the RON2 protein binds to the AMA-1 protein have been identified. It also appears that the formation of the junction and parasitophorous vacuole are molecularly distinct steps in the invasion process.
The invasion process appears to be ATP dependent and may involve a purogenic signalling pathway.
Reticulocyte binding like protein homologue 2a (PfRH2a) is processed both in the schizont as well as during invasion resulting in proteins with different erythrocyte binding properties. It also moves from the rhoptry neck to the moving junction during merozoite invasion. PfRh2a undergoes a cleavage event in the transmembrane region during invasion consistent with activity of the membrane associated PfROM4 protease. Both PfRh2a and PfRh2b bind to red blood cells. The erythrocyte-binding domain lies within a 15 kDa region at the N-terminus of each protein.
A GPI-anchored micronemal antigen (GAMA) appears to be essential in the process of erythrocyte invasion.
PfRh4 binds to a second protein P. falciparum Rh5 interacting protein (PfRipr). PfRipr has a molecular weight of 123 kiloDaltons with 10 epidermal growth factor
-like domains and 87 cysteine
residues distributed along the protein.
The P. falciparum apical sushi protein is the homolog of the P. vivax RON1 protein.
The receptor for the PfRh5 protein appears to be the Ok blood group antigen, basigin
. Blocking access to this protein on the erythrocyte surface appears to inhibit erythrocyte invasion completely. Binding of the Rh5 protein appears to be critically dependent on a single residue within the Rh5 protein.
, S6 and TLP have been implicated in these processes. Heparin
like molecules bound to the surface of the erythrocyte appear to be important in this process which involves the merozoite surface protein 1.
The rhoptery neck proteins (RONs) along with the micronemal AMA1 protein are important in the penetration of the erythrocyte. These form part of the moving junction which initially binds to the erythrocyte surface and is involved in the entry of the parasite into the erythrocyte cytoplasm. The mechanisms involved in this process are still being elucidated. The protein RON8 appears to be central to the binding of parasite to the erythrocyte surface.
Another protein associated with these structure is skeleton-binding protein 1 (SBP1). This protein is invovled in transport of the var gene protein (erythrocyte membrane protein 1) to the erythrocyte surface.
Mutations in the ring exported protein 1 (Rex 1), a protein normally found in Maurer's clefts, reduces transport ofthe var gene products to the erythrocyte surface.
resides inside the parasitophorous vacuole. This is formed during erythrocyte invasion.
The proteins originating in the parasite pass through the membrane of the parasitophorous vacuole and are transported to the cytoplasm or membrane of the erythrocyte. Although this transport mechanism is largely unknown some details have been elucidated. Ingestion of the erythrocyte cytoplasm begins in mid-ring-stage parasites. Host cytoplasm
is internalised via cytostome
-derived invaginations and then concentrated into several acidified peripheral structures. Haemoglobin digestion and haemozoin
formation occur within these vesicle
s. The ring-stage parasites can adopt a deeply invaginated cup shape, but they do not take up haemoglobin via macropinocytosis. As the parasite matures the haemozoin containing compartments coalesce to form a single acidic digestive vacuole (pH 4.5 - 5.5) that is then fed by haemoglobin containing vesicles. Some haemoglobin degradation also occurs in compartments outside the digestive vacuole.
The enzyme phosphatidyl-inositol-3-kinase (PI3K) has been implicated in this process. PI3K is located in vesicular compartments near the membrane and in the food vacuole and is involved in endocytosis from the host and trafficking of hemoglobin in the parasite. Its inhibition with wortmannin
or LY294002
results in entrapment of hemoglobin in vesicles within the parasite cytoplasm preventing its transport to the food vacuole.
The pH of the digestive vacuole is maintained by a V-type H(+)-ATPase.
A signal sequence at the N terminal of proteins targeted to the arasitophorous vacuole has been identified. The signal appears to reside in the 55 amino acids of the N terminal of the protein. There may be a retention signal at the C terminal.
The micronemal protease ROM1 appears to be essential for proper parasitophorous vacuole modification to allow parasite development. This protease is able to cleave the proteins AMA1 and MAEBL.
, and most other members of the phylum
Apicomplexa
, contain an organelle
termed an apicoplast
. The apicoplast
is an essential
plastid
, homologous to a chloroplast
, although the apicoplast
is not photosynthetic. Evolutionarily, it is thought to have derived through secondary endosymbiosis.
The function of the apicoplast
remains to be fully determined, but it appears to be involved in the metabolism of fatty acids, isoprenoids, and heme
.
The apicoplast
contains a 35-kb genome
, which encodes for 30 proteins. Other, nuclear-encoded, proteins are transported into the apicoplast
using a specific signal peptide
. It is estimated that 551, or roughly 10%, of the predicted nuclear-encoded proteins are targeted to the apicoplast
.
As humans do not harbor apicoplasts, this organelle and its constituents are seen as a possible target for antimalarial drugs.
The plastid genome replicates at the late trophozoite stage of the parasite intraerythocytic cycle. It proceeds predominantly via a D-loop/bi-directional ori mechanism with replication ori localized within the inverted repeat region. The process of replication involves a nuclear-encoded DnaJ homolog that binds to the ori site.
The DNA polymerase
involved in the replication of its genome is Pfprex (Klenow-like polymerase). This enzyme has been cloned, expressed and purified. The enzymes is relatively error prone and shows a bias toward T->C mutauons.
The bisynthesis of this organelle is not well understood. Phosphatidylinositol 3-monophosphate has been shown to be involved in its biosynthesis in the apicomplexian Toxoplasma gondii
. It seems likely that this enzyme is involved in the formation of this organells in the Plasmodium species also.
Transport into the apicoplast are not well understood. These proteins has a signal in the N terminal but unlike many other organisms this appears to be a disordered chain rather than a conserved sequence.
Iron-sulphur prosthetic groups are assembled in this organelle. One component (SufB) is encoded in the apicoplast genome and a second (SufC
) is encoded in the nucleus. SufB also exhibits ATPase activity. Other pathways that have been linked to this organelle include biosynthesis of isoprenoid precursors, fatty acids, heme and lipoic acid.
A gene Plasmodium-specific Apicoplast protein for Liver Merozoite formation (PALM) has been shown to be important for merozoite formation. Knock out mutants are unable to release merozoites into the blood from the liver stages. Mutants lacking this gene appear to be able to elicit at least temporary immunity.
The role of the apicoplast in the blood stages has been clarified. Inhibition of isoprenoid precursor biosynthesis with the antibiotic fosmidomycin
(an inhibitor of the enzyme DOXP reductoisomerase) causes delayed death in this parasite. This effect can be overcome with the addition of isopentenyl pyrophosphate
(IPP) to the culture medium. Continued culture in the presence of this agent leads to the loss of the apicoplast genome and these mutants fail to process or localize organelle proteins. These auxotrophs can be grown indefinitely in asexual blood stage culture but are entirely dependent on exogenous IPP for survival.
and the hydrogen ion translocating NADH dehydrogenase
(Complex I, NDH1). The mitochondrion
contains a minimal DNA genome (~6 kilobases) and carries out oxidative phosphorylation in the insect vector stages by using 2-oxoglutarate as an alternative means of entry into the tricarboxylic acid cycle and a single-subunit flavoprotein as an alternative NADH dehydrogenase (NDH2). In the blood stages mitochondrial enzymes are down regulated and parasite energy metabolism relies mainly on glycolysis.
The ATP synthase
is localised to the mitochondrion, is assembled as a large dimeric complex and appears to be essential for in the blood stages of the life cycle. Its function in these stages is not yet clear.
The dicarboxylate-tricarboxylate carrier homolog has been cloned from P. falciparum. This protein may mediate the oxoglutarate-malate
exchange across the inner mitochondrial membrane required for the branched pathway of tricarboxylic acid metabolism.
The ClpQ protease and ClpY ATPase have been cloned. ClpQY function disruption caused hindrance in the parasite growth and maturation of asexual stages of parasites. Features of apoptosis like cell death are also found.
partially permeabilized the parasite's digestive vacuole membrane and that this event appears to precede mitochondrial dysfunction.
.
species have two or three distinct SSU rRNA (18S rRNA) molecules encoded within the genome. These have been divided into types A, S and O. Type A is expressed in the asexual stages; type S in the sexual and type O only in the oocyte. Type O is only known to occur in Plasmodium vivax
at present. The reason for this gene duplication is not known but presumably reflects an adaption to the different environments the parasite lives within.
The Asian simian Plasmodium species - Plasmodium coatneyi, Plasmodium cynomolgi, Plasmodium fragile, Plasmodium inui
, Plasmodium fieldi
, Plasmodium hylobati and Plasmodium simiovale - have a single single S-type-like gene and several A-type-like genes. Phylogenetic analyses has shown that gene duplication events giving rise to A- and S-type-like sequences took place independently at least three times in the Plasmodium evolution.
RNA
helicase
of the DDX6 family, termed DOZI.
The Puf2 gene, a member of the Puf family of transcriptional regulators, has been shown to be involved in gamete formation.
FACT
(facilitates chromatin transcription) is a dimeric complex of two protein
s - SPT16
and SSRP1 - which acts as a histone
chaperone in the (dis)assembly of nucleosome
(and chromatin
) structure during transcription
and DNA replication. It is an essential gene in Plasmodium. Changing its promoter to one expressed only in the blood stages leads to changes in the male gametocytes. The mutant gametocytes have delayed DNA replication and gametocyte formation. Male gamete fertility is strongly reduced. Female gametocytes appear to be normal. When successful fertilization is achieved, the ookinetes generate oocysts that arrest early in development and fail to enter sporogony.
Large holes appear in the cytoskeleton ~35 hours post invasion. This occurs at the same time as the loss of cytoskeletal adaptor proteins that are part of the junctional complex, including α/β-adducin and tropomyosin
. This is followed by the proteolysis of many cytoskeletal proteins during egress at ~48 hours post infection. This later proteolysis is mediated by the erythrocyte's own calpain
-1.
Along with the release from the erythrocyte of the merozoites, the now functionless digestive vacuole is also released. These are can active complement and are rapidly taken up by the polymorph
s. On ingestion the digestive vacuoles induce a vigorous respiratory burst which drives the cells into a state of functional exhaustion, blunting production of reactive oxygen species and microbicidal activity upon challenge with bacterial pathogens.
The serine repeat antigen (SERA) multigene family encode a series of proteins with a putative papain
-like cysteine
protease
motif. One of these SERA5 (120 kiloDaltons) is produced at the late trophozoite/schizont stage. It is secreted together with other SERAs into the parasitophorous vacuole in an infected erythrocyte where it is cleaved into three fragments: an N-terminal domain (47 kDa), a central domain containing putative papain-like cysteine protease motifs (56 kDa) and a C-terminal domain (18 kDa). This N-terminal fragment is then cleaved in turn into two 25 kDa fragments. These fragments become covalently linked to the C-terminal 18 kDa fragment via disulfide bonding and attach to the merozoite surface. The central fragment is further cleaved to 50 kDa and 6 kDa fragments before being shed to the medium. These proteolytic cleavages are carried out by a subtilisin
-like serine
protease called PfSUB1 and the inhibition of this processing, likewise, results in blockade of merozoite release. SERA6 may also be involved in schizont rupture and merozoite release from the erythrocyte.
A protein - gamete egress and sporozoite traversal - has been identified that appears to be involved in the egress of male and female gametes from the erythrocyte. It is also involved in sporozoite migration.
s are known in P. falciparum very little is known about them. Several are cyclin
dependent kinase kinase like kinases (CLK): of these two - the Lammer kinase homologue PfCLK-1 and PfCLK-2 have been cloned. CLKs in other eukaryotes are involved in the regulation of mRNA splicing through phosphorylation of serine
/arginine
-rich proteins. Both are transcribed throughout the asexual blood stages and in gametocytes. PfCLK-1/Lammer possesses two nuclear localization signal sites while PfCLK-2 possesses one of these signal sites upstream of the C-terminal catalytic domains. The two PfCLKs form complexes with proteins with predicted nuclease
, phosphatase
or helicase
functions.
Although the kinases are primarily localized in the parasite nucleus, PfCLK-2 is also present in the cytoplasm. They are important for completion of the asexual replication cycle. Substrates phosphorylated by the PfCLKs include the Sky1p substrate, splicing factor Npl3p, and the plasmodial alternative splicing factor PfASF-1.
Within the genome is a family of four protein kinases (Pfnek-1 to -4) that are related to the NIMA (never-in-mitosis) family of kinases. The members of this latter family play important roles in mitosis
and meiosis
. Pfnek-1 is expressed in asexual parasites and male gametocytes. It is an essential gene for completion of the asexual cycle. The other three (Pfnek-2, -3 and -4) are expressed predominantly in gametocytes.
A mitogen activated protein kinase (MAP kinase) gene is located on is located on chromosome 14. It is predominantly expressed in gametocytes and gametes/zygotes. The protein has 882 amino acid residues and posses a TDY dual phosphorylation site upstream of the highly conserved VATRWYRAPE sequence within subdomain VIII. Within the carboxyl-terminal segment the protein contains an unusually large and highly charged domain. This region includes two repetitive sequences of either a tetrapeptide or octapeptide motif.
A subgroup of cyclin-dependent kinases (CDK) including crk-5 have an activation loop that contains a novel Proline
-Threonine
-x-Cytosine
motif which is absent from all known CDKs outside the Apicomplexa.
The plasmodial mitogen-activated protein kinase kinase Pfnek3 has both serine
/threonine
and tyrosine kinase
activities.
homolog. The falcipains belong to the papain
family of enzymes. Falcipain-1 appears to be important in the development of the oocysts in the mosquito. Falcipain-2 is involved in the hydrolysis of haemoglobin and appears to be a non essential gene. Falcipain-3 appears to be an essential gene but its function has yet to be firmly established. Dipeptidyl aminopeptidase 1 is found in the food vacuole and is also an essential gene. Dipeptidyl aminopeptidase 3 appears to be involved in the release of the merozoites from the erythrocyte. SERA-5 and SERA-6 appear to be essential genes: SERA-5 also seems to be involved with egress of the merozoites.
There are at least 2 essential metallopeptidases encoded in the genome - PfA-M1 and Pf-LAP. Specific inhibition of PfA-M1 causes swelling of the parasite digestive vacuole and prevented proteolysis of haemoglobin derived oligopeptides. This inhibition is lethal to the parasite probably by starvation. Inhibition of Pf-LAP is lethal to parasites early in the life cycle, prior to the onset of haemoglobin degradation suggesting a different role for this enzyme.
s known as plasmepsins are involved in the degradation of hemoglobin
by Plasmodium falciparum.
The histo-aspartic protease (HAP) has been crystalised. This protein has high sequence similarity to pepsin
-like aspartic
proteases, but one of the two catalytic aspartates, Asp32, is replaced in this enzyme by a histidine
residue. The propeptide interacts with the C-terminal domain of the enzyme, forcing the N- and C- terminal domains apart. This mechanically separates His32 and Asp215 and prevents formation of the mature active site. This mechanism is similar to those of other proplasmepsins. The enzyme has a number of unique features and may be a useful drug target.
A SUMO
specific protease PfSENP1 (PFL1635w) has been identified in the genome but its importance if any is not known
The parasite is capable of making use of the erythrocyte's own enzymes. The enzymes PAK1
and MEK1 neither of which are encoded in the Plasmodium genome have been shown to be phosphorylated and activated during the course of infection' In vitro work has shown that inhibition of these enzymes is fatal to the parasite.
The intracellular concentration of chloride ions has been estimated to be 48 milliMolar. It appears to actively import using ATP
both hydrogen
ion
s and chloride
ions in a linked fashion via a DIDS sensitive transporter in the cytoplasmic membrane.
One difficulty the parasite has in acquiring nutrients from the cytoplasm is the presence of phosphate groups on these molecules. It appears to have overcome this by secreting an acid phosphatase (glideosome-associated protein 50 - GAP50 ) into the cytoplasm that is then taken up into the food vacuole.
The parasite is unable to synthesize purines (including adenosine, hypoxanthine
and adenine
) and must take these up from the host. Three purine transporters have been studied: the human equilibrative nucleoside transporter
(hENT1), the human facilitative nucleobase transporter (hFNT1) and the parasite-induced new permeation pathway (NPP). The bulk of transport is facilitated by host's own transporters rather than through the NPP. Hypoxanthine and adenine were transported mainly through the hFNT1 pathway whereas adenosine entered predominantly through the hENT1 system. The rate of purine uptake in infected cells was approximately twice that of uninfected erythrocytes. The rate of adenosine uptake was greater than the rate of hypoxanthine uptake in infected human red blood cells. Furosemide
inhibits the transport of purine bases through the hFNT1.
The parasite possesses its own Equilibrative Nucleoside Transporter 1. All members of this protein family have 11 transmembrane segments. The gene product is located in the parasite's plasma membrane and knock out mutants have shown that this is an essential gene at least at physiological concentrations. In the 11th transmembrane segment two mutations have been shown to affect its activity: a phenylalanine
(Phe) to leucine
(Leu) at residue 394 (F394L) via cytosine
(C) or uracil
(U) to adenosine (A) or guanine
(G) at the third codon position and a cysteine
(Cys) to glycine
(Gly) mutation at either glycine in a conserved glycine-X-X-glycine motif (where X is any amino acid) via a cytosine to uracil at the second codon position. Additional work suggests that the 11th transmembrane segment is largely alpha helical. It has been suggested that this transmembrane segment may be the actual purine transport channel.
The parasite has an absolute requirement for isoleucine
- an amino acid
absent from human haemoglobin. A saturable neutral amino acid (methionine
, leucine
, isoleucine) transporter appears to be encoded by the parasite and this protein functions in the infected erythrocyte membrane.
The clag3 on chromosome 3 appear to be involved in anion transport rather than in cell adherence as originally thought.
The P. falciparum NA+/H+ exchanger (PfNHE1) is located on chromosome 13 (gene PF13_0019).
Two folate transporters (PfFT1 and PfTF2) have been cloned. Substrates include folic acid
, folinic acid
, the folate precursor pABA and the human folate catabolite pABAG(n). 5-methyl tetrahydofolate is not transported by PfFT1 and only poorly by PfFT2. The activity of both transporters may be inhibited by probenecid
or methotrexate
.
An intracellular purine
permease
(PfNT2) has been shown to be localised to the endoplasmic reticulum
. This protein is a member of the equilibrative nucleoside transporter family.
The clag3 gene family encode a parasite ion channel known as the plasmodial surface anion channel. Its activation appears to involve an intracellular domain.
- I and II. The first form (I) is present in significantly greater quantities. Actin II appears to be essential for the process of exflagellation. Deletion of this gene results in viable asexual stages. During the formation of the male gametes actin I is found initially in both the nucleus and the cytoplasm. After activation it is found only in the cytoplasm. In actin II deletion mutants actin I remains in both the nucleus and the cytoplasm after activation. Morphologically in the actin II mutants male gametocyte DNA was replicates normally and axonemes are assembled but egress from the host cell is inhibited and axoneme motility is abolished.
Two proteins P. falciparum actin-depolymerizing factor 1 (PfADF1) and P. falciparum actin-depolymerizing factor 2 (PfADF2) are involved in the polymerisation of actin. PfADF1 has ben crystalised and despite having significant differences from other proteins with similar function it is capable of severing actin filaments. PfADF2, like canonical ADF proteins but unlike ADF1, binds to both globular and filamentous actin, severing the filaments and inducing nucleotide exchange on the actin monomer. The crystal structure of PfADF1 shows major differences from the ADF consensus, explaining the lack of F-actin binding. PfADF2 structurally resembles the canonical members of the ADF/cofilin family.
The actins found In Plasmodium and in Toxoplasma are divergent both in sequence and function and only form short, unstable filaments in contrast to the stability of conventional actin filaments. This inherent instability of parasite's actin filaments is a critical adaptation for their gliding motility.
Actin is involved in the expression of the var genes. The var introns interact with an 18 base pair nuclear protein binding element which recruits actin and repositions the var DNA from a transcriptionally repressive to a transcriptionally active perinuclear compartment.
is bound to the surface of P. falciparum and several other pathogens. In this location it binds plasminogen which is thought to function in the degradation of the extracellular matrix surrounding the targeted host cell, thereby facilitating pathogen invasion.
The ETRAMP family is characterized by a predicted signal peptide
, a short lysine
rich stretch, an internal transmembrane domain and a highly charged C-terminal region of variable length. The highly charged terminal region appears to be involved in protein-protein interactions. The gene ETRAMP 10.3 has been shown to be expressed in the liver, sporozoites and blood stages. Within the liver and blood stages it is localized to the parasitophorous vacuole membrane. It is also exported to the erythrocyte during the blood stages. It appears to be an essential gene in the blood stages.
The receptor for the attachment protein PfRh4 has been identified as complement receptor 1
.
The TRAP like protein is invovled in cell motility.
The circumsporozoite- and thrombospondin-related adhesive protein (CTRP) is a modular multidomain protein containing six tandem von Willebrand factor
A like domains and seven tandem thrombospondin
type I repeat-like domains. The A domains of CTRP are critical for ookinete gliding motility and oocyst formation. The thrombospondin domains are fully redundant.
The cell-traversal protein for ookinetes and sporozoites (CelTOS) is a protein involved in the invasion of both vertebrate and insect host cells.
Merozoite surface protein 7 appears to enhance the virulence of the parasite at least in the rodent.
Merozoite surface protein
2 is one of the most abundant proteins on the surface of merozoites, is intrinsically unstructured and forms amyloid
-like fibrils in solution.
The mature parasite-infected erythrocyte surface antigen (MESA) is exported to the erythrocyte cytoplasm where it binds to the N-terminal 30 kiloDalton domain of the erythrocyte protein 4.1R via a 19-residue sequence. This sequence is also found in a number of other proteins in the parasite. Their role in remodeling of the erythrocyte are still under investigation.
The proteins Pf12, Pf34, Pf92 and Pf38 are associated with detergent resistant membrane microdomains through glycosylphosphatidylinositol anchor sequences. These microdomains are considered organizing centers for the assembly of molecules implicated in cell signaling.
Positive diversifying selection is present in clag2, clag8 and clag9 but not in clag3.1 and clag3.2.
to other proteins as part of post translational processing is widespread in most eukaryotes. This is also the case with P. falciparum where this process occurs at all stages of the asexual life cycle.
A homolog of the DEAD box
(Asparagine
-Glutamate-Alanine
-Asparagine) RNA
helicase
DDX19 (Dbp5) has been cloned from the P. falciparum genome and has been termed PfD66. This protein has intrinsic nucleic acid dependent ATPase
and RNA binding activities and ATP dependent bipolar DNA
and RNA unwinding activity.
SAP1 has been shown to be involved in the post transcriptional control of liver
stage genes.
The protein PfMyb1 is a transcription factor belonging to the tryptophan
cluster family. Inhibition of this gene reduces growth by ~40% with the mortality being concentrated at the trophozoite-schizont interface.
s 40 (hsp40) have been predicted from the sequenced genome. Only one is predicted to be a cytosolic canonical Hsp40 capable of interacting with the major cytosolic Hsp70 an interaction that has been confirmed experimentally.
PfGECO is a type IV heat shock protein 40 expressed in gametocyte stages I to IV and is exported to the erythrocyte cytoplasm. This gene appears to be non essential.
Heat shock proteins Hsp70 and Hsp90 are both expressed in P. falciparum. They are linked by an essential adaptor protein known as the Hsp70-Hsp90 organising protein (Hop). This protein co-localises with PfHsp70 and PfHsp90 at the trophozoite stage and forms a complex with them.
The protein Aha1 interacts with HSP90.
(PfAdoMetDC/ODC). On the secondary structure level PfAdoMetDC is similar to that of the human protein. This bifunctional enzyme ensure coordination decarboxylated AdoMet and putrescine
for the subsequent synthesis of spermidine
.
P. falciparum contains both cytosolic and mitochondrial serine
hydroxymethyltransferase isoforms.
The first two reactions of the pentose phosphate pathway
in P. falciparum are catalysed by a single bifunctional enzyme - glucose 6-phosphate dehydrogenase 6-phosphogluconolactonase
. This is distinct from the case in humans where the enzymes are separate. In animal
s this pathway is usually found in the cytosol while in plant
s it is found in the plastid
s. The location of this reaction is not currently known in P. falciparum.
Fusions between these two enzymes (glucose 6-phosphate dehydrogenase and 6-phosphogluconolactonase
) have also been reported in chordates. The chordate fusion differs in its orientation to that in Plasmodium (in Plasmodium the 6-phosphogluconolactone is found at the N-terminus of the glucose 6-phosphate dehydrogenase protein) indicating that at least two separate fusion events have occurred. The metazoan fusion appears to have occurred near the bases of the metazoan and apicomplexa
n lineages. This fusion event was not found in any of the three sequenced Cryptosporidium
genomes. It was not found in Perkinsus marinus
or in either of the ciliate (Paramecium tetraurelia and Tetrahymena thermophila) genomes. More data will be needed to estimate the timing of this fusion event.
Only one of the one of the two metazoan paralogs of glucose 6-phosphate dehydrogenase is fused indicating that the fusion occurred after a duplication event. This duplication event occurred in an ancestor of the choanoflagellate
s and metazoans. Another fusion event between these enzymes occurred in an ancestor of the protozoan parasites Trichomonas
and Giardia lamblia
. In Giardia the proteins are fused in opposite orientations. A third fusion event occurred between glucose 6-phosphate dehydrogenase with phosphogluconate dehydrogenase
in a diatom
species (Phaeodactylum tricornutum
).
Phosphoinositide-specific phospholipase
C (PI-PLC) is a major regulator of calcium
-dependent signal transduction usually by liberation of calcium from intracellular stores through the action of its product, inositol-(1,4,5)-trisphosphate. These genes are found in P. falciparum and appear to be essential. The genes are twice as long as their mammalian counterparts and belong to the delta class of phospholipase C proteins.
The mechanism of action of the triose phosphate isomerase enzyme has been investigated in some detail. The conserved glutamic acid
residue at position 97 is involved in the catalytic proton transfer. Modification of this residue may reduce the rate of catalysis by 9000 fold.
There are two translation elongation factor g
proteins encoded in the genome. One is located in the mitochondrion and the second in the plastid. Both appear to be inhibitable with fusidic acid
The type II NADH:ubiquinone oxidoreductase
has been shown to be redundant in the blood forms but to be essential in the mosquito midgut.
Chorismate synthase
(CS) catalyses the seventh and final step of the shikimate pathway. P. falciparum chorismate synthase (PfCS) is unique in terms of enzymatic behavior, cellular localization and in having two additional amino acid inserts compared to any other CS.
Polynucleotide kinase/phosphatase (PNKP) is a bifunctional enzyme that can phosphorylate the 5'-OH termini and dephosphorylate the 3'-phosphate termini of DNA
. It is a DNA repair enzyme involved in the processing of strand break termini, which permits subsequent repair proteins to replace missing nucleotides and rejoin broken strands. A P. falciparum gene encoding a protein with 24% homology to human PNKP has been cloned. This enzyme dephosphorylates single-stranded substrates or double-stranded substrates with a short 3'-single-stranded overhang, but not double-stranded substrates that mimicked single-strand breaks.
Sir2A is a member of the sirtuin
family of nicotinamide adenine dinucleotide
dependent deacetylases. In P. falciparum it has been has been shown to regulate the expression of surface antigens to evade the detection by host immune surveillance. While it is a poor deacetylator of histone
s it also catalyzes the hydrolysis of medium and long chain fatty acyl groups from lysine
residues. Proteins are present in P. falciparum with these modifications and these can be removed by can be removed by PfSir2A in vitro. This suggests that this may be its role rather than the deacetylation of histones.
The telomerase
(tert) is a large protein (2518 codons) and has a predicted molecular weight of ~280 kiloDaltons. It has the usual telomerase specific motifs within the N-terminal half of the protein (GQ/N, CP, QFP and T) and reverse transcriptase (RT) specific motifs in the C-terminal half. The N-terminal half is required for efficient binding of the RNA template, defining the 5′ RNA template boundary, multimerization and interactions with associated proteins. The RT domain is essential for the catalytic activity. The protein contains several nuclear localization signals and is found in the nucleolus
.
A histone deacetylase
(HDAC1) has been cloned. The protein has 449 amino acid residues and localises to the nucleus. Its molecular weight is 50 kiloDaltons and it is predominantly expressed in mature asexual blood stages and in gametocytes.
A phosphatidylserine decarboxylase
has been cloned from the parasite Plasmodium knowlesi
. It seems highly probably this enzyme is also found in P. falciparum.
Tyrosyl tRNA synthetase is secreted by the parasite into the cytoplasm
of the infected erythrocyte. On lysis of the erythrocyte it is released into the blood stream where it is pro inflamatory. It is specifically bound by and taken up by host macrophages and leads to enhanced secretion of the cytokine
s tumor necrosis factor-alpha and interleukin 6
. This interaction also increases the adherence linked host endothelial receptors ICAM-1
and VCAM-1
.
The MutL homolog (MLH) - part of the DNA mismatch repair
system - has been cloned. MLH possess ATPase
and endonuclease
activities. Its' expression is maximal in the schizont stage.
Both the 2-C-methyl-d-erythritol-4-phosphate and shikimate pathways are functional in P. falciparum and vitamin E
biosynthesis occurs.
Membrane biogenesis in this organism involves the enzyme phosphoethanolamine methyltransferase which catalyses the methylation of phosphoethanolamine to phosphocholine
. This pathway is found in plant
s and nematode
s but not in humans. The enzymes in P. falciparum is a multi-functional unlike that of plants and nematodes. The enzyme from P. falciparum has been cloned and its structure solved.
A homolog of the inhibitor of protein phosphatase 1
has been cloned. This gene is essential for survival and appears to be localised to the nucleus. A conserved 41- Lysine
-Valine
-Valine-Arginine
-Tryptophan
- 45 motif is essential for its inhibition activity.
(clone 3D7) was fully sequenced in 2002. The parasite has a 23 megabase genome, divided into 14 chromosomes. The genome codes for approximately 5,300 genes. About 60% of the putative proteins have little or no similarity to proteins in other organisms, and thus currently have no functional assignment. It is estimated 52.6% of the genome is a coding region, with 53.9% of the putative genes containing at least one intron.
gut.
genome has an AT content of roughly 80.6%. Within the intron and intergenic regions, this AT composition rises to roughly 90%. The putative exons contain an AT content of 76.3%. The parasite's AT content is very high in comparison to other organisms. For example, the entire genomes of Saccharomyces cerevisiae
and Arabidopsis thaliana
have AT contents of 62% and 65%, respectively.
at the -1 position resulted in 2.5-fold higher reporter activity compared to wild type.
and 54 candidate recombination hotspots have been identified. The centromere
s are found in chromosome regions largely devoid of recombination activity like other organisms. Within the hotspots a number of motifs were enriched including a 12 base pair G/C-rich motif with 3 base pair periodicity that may interact with a protein containing 11 predicted zinc finger
arrays.
Subtelomeric regions in general are low in gene density, low in transcription, low in recombination, late replicating, are involved in protecting the end from degradation and end-to-end fusions and in completing replication. The subtelomeric repeats can rescue chromosome ends when telomerase
fails, buffer subtelomerically located genes against transcriptional silencing and protect the genome from deleterious rearrangements due to ectopic recombination. They may also be involved in fillers for increasing chromosome size to some minimum threshold level necessary for chromosome stability; act as barriers against transcriptional silencing; provide a location for the adaptive amplification of genes; and be involved in secondary mechanism of telomere maintenance via recombination when telomerase activity is absent. The repressive histone
3 lysine 9 tri-methylation mark and heterochromatin protein 1
are found throughout the TAS region and adjacent gene families on all chromosomes. These heterochromatic marks are important in telomere proximal gene silencing.
In parasitic species genes involved in antigenic variation are commonly located in these regions.
The chromsomes of P. falciparum conform to this basic pattern with the ends of the chromosomes consist of a stretch of telomeric GGGTT(T/C)A repeats with an average size of 1.2 kilobases (kb). This is followed by an extensive 20 to 40 kb TAS domain. These show a high degree of conservation within the genome and contain significant amounts of repeated structure. Telomere repeats are followed by a mosaic of six distinct telomere associated repetitive elements (TAREs 1-6) which are always found in the same order but vary in size.
Many genes involved in antigenic variation are located in the subtelomeric regions of the chromosomes. These are divided into the var, rif, and stevor families. Within the genome, there are 59 var, 149 rif, and 28 stevor genes along with multiple pseudogenes and truncations.
The chromosome ends form clusters of 4–7 telomeres that localize around the nuclear periphery. Within this location the telomeric areas undergo frequent recombination which seems to increases antigenic variation.
Transcription in P. falciparum appears to have significant differences from that found in most other eukaryotes examined to date with the chromatin undergoing dramatic unheavals during the cell cycle.
A transcriptome analysis has been conducted on the intraerythrocytic development cycle of P. falciparum
. Roughly 60% of the genome is transcriptionally active during this portion of the parasite's life cycle. Whereas many genes appear to have stable mRNA levels throughout the cycle, many of the genes are transcriptionally regulated in a continuous cascade.
The transition from early trophozoite to trophozoite to schizont correlates with the ordered induction of genes related to transcription/translation machinery, metabolic synthesis, energy metabolism, DNA replication, protein degradation, plastid functions, merozoite invasion, and motility.
Closely adjacent genes along the chromosome do not exhibit common transcription characteristics. Thus, genes are likely individually regulated along the parasite chromosome.
Conversely, the apicoplast genome is polycistronic and most of its genes are coexpressed during the intraerythrocytic development cycle.
, and roughly 60% share little or no similarity to proteins in other organisms and thus are without functional assignment. Of the predicted proteins, 31% contain at least one transmembrane domain, and 17.3% have a signal peptide or signal anchor.
It is estimated that 10.4% of the proteome
is targeted to the apicoplast
.
It is estimated that 4.7% of the proteome
is targeted to the mitochondria.
The parasite has different subsets of its proteome expressed during various stages of its developmental cycle. In one study, of the 2,415 proteins were identified in four stages(sporozoite, merozoite, trophozoite, gametocyte), representing 46% of the theoretical number of proteins. Only 6% of the proteins were found in all of the four stages. Of the proteins found, 51% were annotated as hypothetical proteins.
Merozoites contained high levels of cell recognition and invasion proteins. Trophozoites contained proteins implicated in erythrocyte remodeling and hemoglobin
digestion. Gametocytes contained high amounts of gametocyte-specific transcription factors and cell cycle/DNA processing proteins. The gametocytes had low levels of polymorphic surface antigens. Sporozoites contained large amounts of proteins related to invasion, as well as members of the var and rif families.
have yet to be fully elucidated, the presence and components of many can be predicted through genomic analysis.
as a food source. The protein is broken down into peptides, and the heme group is released and detoxified by biocrystallization
in the form of hemozoin.
Heme
biosynthesis by the parasite has been reported.
. This competition may be fatal to the parasite. To detoxify the ferriprotoporphyrin IX the parasite polymerizes hematin to hemozoin which is the characteristic malaria pigment.
Artemisinin
an anti malarial agent appears to require haemoglobin digestion for its activity. Inhibition of hemoglobinase activity with cysteine protease inhibitors, knockout of falcipain 2 or deprivation of host cell lysate reduce the activity of the drug against the parasite.
Haemozoin has multiple effects on the hosts immune system including increasing the level of matrix metalloproteinase-9.
.
Genes encoding for the TCA cycle enzymes are present in the genome, but it is unclear whether the TCA cycle is used for oxidation of glycolytic products to be used for energy production, or for metabolite intermediate biosynthesis. It has been hypothesized that the main function of the TCA cycle in P. falciparum
is for production of succinyl-CoA, to be used in heme biosynthesis.
Genes for nearly all of the pentose phosphate pathway
enzymes have been identified from the genome sequence.
. This is supported by the fact that genomic analysis has found no enzymes necessary for amino acid
biosynthesis, except for glycine-serine, cysteine-alanine, aspartate-asparagine, proline-ornithine, and glutamine-glutamate interconversions
.
Much of this digestion occurs within the digestive vacuole. Multiple enzymes are involved in this process including four distinct plasmepsins (aspartic acid proteases).
is a major and essential membrane phospholipid in the parasite. Its synthesis occurs via the CDP-choline and the serine decarboxylase phosphoethanolamine methylation pathways. The substrates of these pathways are the host's choline
, serine
and fatty acids. Both pathways share the final two steps catalyzed by two essential enzymes CTP:phosphocholine cytidylyltransferase and choline-phosphate transferase.
β-hydroxyacyl-acyl carrier protein dehydratase catalyzes the third and important reaction of the fatty acid elongation cycle. The P. falciparum gene has been cloned and the crystal structure of the enzyme solved.
is unable to biosynthesize purines. Instead, the parasite is able to transport and interconvert host purines.
Conversely, the parasite can produce pyrimidines de novo using glutamine, bicarbonate, and aspartate.
The proteins encoded by the var genes are ultimately transported to the erythrocyte membrane and cause the infected erythrocytes to adhere to host endothelial receptors. Due to transcriptional switching between var genes, antigenic variation occurs which enables immune evasion by the parasite.
The protein PfSir2 associates with promoter regions of silenced genes involved in antigenic variation.
Rifin protein are ultimately transported to the erythrocyte membrane. The function of these proteins is currently unknown.
These proteins appear to affect membrane deformability.
Bergman et al (2003) Myosin A tail domain interacting protein (MTIP) localizes to the inner membrane complex of Plasmodium sporozoites. J Cell Sci 116 (1) 39–49
Baum et al (2005) Invasion by P. falciparum merozoites suggests a hierarchy of molecular interactions. PLoS Pathog 1(4) e37
Bosch et al (2007) The closed MTIP-myosin A-tail complex from the malaria parasite invasion machinery. J Mol Biol 372(1) 77–88
Bosch et al (2007) Aldolase provides an unusual binding site for thrombospondin-related anonymous protein in the invasion machinery of the malaria parasite. Proc Natl Acad Sci USA 104(17) 7015–20
Daher & Soldati-Favre (2009) Mechanisms controlling glideosome function in apicomplexans. Cur Opin Micro 12 (4) 408–414
Sibley (2010) How apicomplexan parasites move in and out of cells. Cur Opin Biotech 21(5) 592–598
Plasmodium falciparum
Plasmodium falciparum is a protozoan parasite, one of the species of Plasmodium that cause malaria in humans. It is transmitted by the female Anopheles mosquito. Malaria caused by this species is the most dangerous form of malaria, with the highest rates of complications and mortality...
has been the focus of much research due to it being the causative agent of malaria
Malaria
Malaria is a mosquito-borne infectious disease of humans and other animals caused by eukaryotic protists of the genus Plasmodium. The disease results from the multiplication of Plasmodium parasites within red blood cells, causing symptoms that typically include fever and headache, in severe cases...
. This article describes some of the recent findings surrounding the unique biology of this organism.
Life cycle
Plasmodium falciparumPlasmodium falciparum
Plasmodium falciparum is a protozoan parasite, one of the species of Plasmodium that cause malaria in humans. It is transmitted by the female Anopheles mosquito. Malaria caused by this species is the most dangerous form of malaria, with the highest rates of complications and mortality...
has a complicated life-cycle, requiring both a human and a mosquito
Mosquito
Mosquitoes are members of a family of nematocerid flies: the Culicidae . The word Mosquito is from the Spanish and Portuguese for little fly...
host, and differentiating multiple times during its transmission/infection process.
Human infection
P. falciparumPlasmodium falciparum
Plasmodium falciparum is a protozoan parasite, one of the species of Plasmodium that cause malaria in humans. It is transmitted by the female Anopheles mosquito. Malaria caused by this species is the most dangerous form of malaria, with the highest rates of complications and mortality...
is transmitted to humans by the females of the Anopheles
Anopheles
Anopheles is a genus of mosquito. There are approximately 460 recognized species: while over 100 can transmit human malaria, only 30–40 commonly transmit parasites of the genus Plasmodium, which cause malaria in humans in endemic areas...
species of mosquito
Mosquito
Mosquitoes are members of a family of nematocerid flies: the Culicidae . The word Mosquito is from the Spanish and Portuguese for little fly...
. There are about 460 species of Anopheles
Anopheles
Anopheles is a genus of mosquito. There are approximately 460 recognized species: while over 100 can transmit human malaria, only 30–40 commonly transmit parasites of the genus Plasmodium, which cause malaria in humans in endemic areas...
mosquito
Mosquito
Mosquitoes are members of a family of nematocerid flies: the Culicidae . The word Mosquito is from the Spanish and Portuguese for little fly...
, but only 68 transmit malaria. Anopheles gambiae
Anopheles gambiae
Anopheles gambiae is a complex of at least seven morphologically distinguishable species of mosquitoes in the genus Anopheles. This complex was recognised in the 1960s and includes the most important vectors of malaria in sub-Saharan Africa and the most efficient malaria vectors known.This species...
, found in Africa, is one of the best malaria vectors. It is long-lived, prefers feeding on humans, and lives in areas near human habitation.
Prior to transmission, Plasmodium falciparum
Plasmodium falciparum
Plasmodium falciparum is a protozoan parasite, one of the species of Plasmodium that cause malaria in humans. It is transmitted by the female Anopheles mosquito. Malaria caused by this species is the most dangerous form of malaria, with the highest rates of complications and mortality...
resides within the salivary gland of the mosquito
Mosquito
Mosquitoes are members of a family of nematocerid flies: the Culicidae . The word Mosquito is from the Spanish and Portuguese for little fly...
. The parasite is in its sporozoite stage at this point. As the mosquito
Mosquito
Mosquitoes are members of a family of nematocerid flies: the Culicidae . The word Mosquito is from the Spanish and Portuguese for little fly...
takes its blood meal, it injects a small amount of saliva into the skin wound. The saliva contains antihemostatic and anti-inflammatory enzymes that disrupt the clotting process and inhibit the pain reaction. Typically, each infected bite contains 5-200 sporozoites which proceed to infect the human vector. Once in the human bloodstream, the sporozoites only circulate for a matter of minutes before infecting liver cells.
Liver stage
After circulating in the bloodstream, the P. falciparumPlasmodium falciparum
Plasmodium falciparum is a protozoan parasite, one of the species of Plasmodium that cause malaria in humans. It is transmitted by the female Anopheles mosquito. Malaria caused by this species is the most dangerous form of malaria, with the highest rates of complications and mortality...
sporozoites enter hepatocytes. At this point, the parasite loses its apical complex and surface coat, and transforms into a trophozoite. Within the parasitophorous vacuole of the hepatocyte
Hepatocyte
A hepatocyte is a cell of the main tissue of the liver. Hepatocytes make up 70-80% of the liver's cytoplasmic mass.These cells are involved in:* Protein synthesis* Protein storage* Transformation of carbohydrates...
, P. falciparum
Plasmodium falciparum
Plasmodium falciparum is a protozoan parasite, one of the species of Plasmodium that cause malaria in humans. It is transmitted by the female Anopheles mosquito. Malaria caused by this species is the most dangerous form of malaria, with the highest rates of complications and mortality...
undergoes schizogonic development. In this stage, nucleus divides multiple times with a concomitant increase in cell size, but without cell segmentation. This exoerythrocytic schizogony stage of P. falciparum
Plasmodium falciparum
Plasmodium falciparum is a protozoan parasite, one of the species of Plasmodium that cause malaria in humans. It is transmitted by the female Anopheles mosquito. Malaria caused by this species is the most dangerous form of malaria, with the highest rates of complications and mortality...
has a minimum duration of roughly 5.5 days. After segmentation, the parasite cells are differentiated into merozoites.
After maturation, the merozoites are released from the hepatocytes and enter the erythrocytic portion of their life-cycle. Note that these cells do not reinfect hepatocytes.
Plasmodium possess only a single pyruvate dehydrogenase
Pyruvate dehydrogenase
Pyruvate dehydrogenase complex is a complex of three enzymes that transform pyruvate into acetyl-CoA by a process called pyruvate decarboxylation. Acetyl-CoA may then be used in the citric acid cycle to carry out cellular respiration, and this complex links the glycolysis metabolic pathway to the...
enzyme (PDH) complex. This is localized to the plastid-like organelle known as the apicoplast
Apicoplast
An apicoplast is a derived non-photosynthetic plastid found in most Apicomplexa, including malaria parasites such as Plasmodium falciparum, but not in others such as Cryptosporidium. It originated from an algae through secondary endosymbiosis...
. Unlike most eukaryotes, Plasmodium lacks a mitochondrial PDH. The PDH catalyzes the conversion of pyruvate to acetyl-CoA
Acetyl-CoA
Acetyl coenzyme A or acetyl-CoA is an important molecule in metabolism, used in many biochemical reactions. Its main function is to convey the carbon atoms within the acetyl group to the citric acid cycle to be oxidized for energy production. In chemical structure, acetyl-CoA is the thioester...
, an important precursor for the tricarboxylic acid cycle and type II fatty acid
Fatty acid
In chemistry, especially biochemistry, a fatty acid is a carboxylic acid with a long unbranched aliphatic tail , which is either saturated or unsaturated. Most naturally occurring fatty acids have a chain of an even number of carbon atoms, from 4 to 28. Fatty acids are usually derived from...
synthesis.
The process of maturation within the liver is still being investigated. In the species Plasmodium yoelli the exit from the liver appears to be dependent on type II fatty acid synthesis. Deletions in either the pyruvate dehydrogenase E1 alpha and E3 subunits produce a phenotype similar to that found in mutants of the type II fatty acid synthesis pathway. These mutants appear normal in blood stage development, mosquito stage development and early liver stage development but fail to exit the liver cells.
Plasmodium is unable to synthesize sterol
Sterol
Sterols, also known as steroid alcohols, are a subgroup of the steroids and an important class of organic molecules. They occur naturally in plants, animals, and fungi, with the most familiar type of animal sterol being cholesterol...
s they must obtain these from the host. However manipulation of cholestoerol metabolism does not impede the development of the merozoites.
Invasion of the liver cell chances the properties of the cell itself. The cell membrane becomes rougher and the cell itself becomes significantly stiffer. The mechanisms of these changes is currently unknown.
Erythrocytic stage
After release from the hepatocytes, the merozoites enter the bloodstream prior to infecting red blood cells. At this point, the merozoites are roughly 1.5 μm in length and 1 μm in diameter, and use the apicomplexan invasion organelles (apical complex, pellicle and surface coat) to recognize and enter the host erythrocyte.
Unlike species in the genus Toxoplasma which have multiple rhopteries
Rhoptry
A rhoptry is a specialized secretory organelle. They are club-shaped organelles connected by thin necks to the extreme apical pole of the parasite. These organelles, like micronemes, are characteristic of the motile stages of Apicomplexa protozoans. They can vary in number and shape and contain...
, Plasmodium species typically only have two. These may be referred in the literature to as the 'paired organ'.
The mechanism of cell attachment and invasion are under active investigation. Several molecules have been implicated in this process. Two of these are the genes PfRh2a and PfRh2b. These are large proteins of about 3200 amino acids in length. They differ only in the last 500 amino acids of the C terminal and have clearly arisen by a process of gene duplication and mutation. The 500 amino acid region includes an ectodomain, a transmembrane domain and a cytoplasmic domain. PfRh2b is essential for a well-defined invasion pathway while PfRh2a is not required or sufficient for this pathway. It has been shown that the reason for this difference lies in the cytoplasmic domain.
The parasite first binds to the erythrocyte in a random orientation. It then reorients such that the apical complex is in proximity to the erythrocyte membrane. A tight junction is formed between the parasite and erythrocyte. As it enters the red blood cell, the parasite forms a parasitophorous vesicle, to allow for its development inside the erythrocyte.
Five separate invasion paths have been delineated. The most common pathway is neuraminidase
Neuraminidase
Neuraminidase enzymes are glycoside hydrolase enzymes that cleave the glycosidic linkages of neuraminic acids. Neuraminidase enzymes are a large family, found in a range of organisms. The most commonly known neuraminidase is the viral neuraminidase, a drug target for the prevention of the spread...
resistant, trypsin
Trypsin
Trypsin is a serine protease found in the digestive system of many vertebrates, where it hydrolyses proteins. Trypsin is produced in the pancreas as the inactive proenzyme trypsinogen. Trypsin cleaves peptide chains mainly at the carboxyl side of the amino acids lysine or arginine, except when...
sensitive and chymotrypsin
Chymotrypsin
Chymotrypsin is a digestive enzyme that can perform proteolysis. Chymotrypsin preferentially cleaves peptide amide bonds where the carboxyl side of the amide bond is a tyrosine, tryptophan, or phenylalanine. These amino acids contain an aromatic ring in their sidechain that fits into a...
resistant invasion. Some parasites have a neuraminidase- and trypsin-sensitive phenotype indicating a dependence on the erythrocyte binding antigen 175/glycophorin A
GYPA
Glycophorin A , also known as GYPA, is a protein which in humans is encoded by the GYPA gene. GYPA has also recently been designated CD235a .- Function :...
pathway(s). Most isolates appear to be dependent on a trypsin sensitive pathway.
The mechanics of the binding process have been partly elucidated. The low potassium levels in the blood (3.5-5.0 millimoles per liter) activates a phospholipase C enzyme. This enzyme initiates a pathway that leads to a rise in the intracellular calcium. This rise in calcium triggers secretion of microneme proteins including the 175 kD erythrocyte binding antigen and apical membrane antigen 1
Apical membrane antigen 1
In molecular biology, apical membrane antigen 1 is a novel antigen of Plasmodium falciparum which has been cloned. It contains a hydrophobic domain typical of an integral membrane protein. The antigen is designated apical membrane antigen 1 by virtue of appearing to be located in the apical complex...
to the merozoite surface where they can bind to erythrocyte surface proteins including glycophorin A
GYPA
Glycophorin A , also known as GYPA, is a protein which in humans is encoded by the GYPA gene. GYPA has also recently been designated CD235a .- Function :...
.
It appears that the merozoite surface protein 1 (MSP1) binds to heparin
Heparin
Heparin , also known as unfractionated heparin, a highly sulfated glycosaminoglycan, is widely used as an injectable anticoagulant, and has the highest negative charge density of any known biological molecule...
like molecules on the surface of the erythrocyte and ththat is binding is an essential step in the invasion process.
The protein PfRON2 via the C-terminal as well as the central cysteine-rich domain interacts with PfAMA1. This protein also appears to be involved in erythrocyte invasion.
Trophozoite
After invading the erythrocyte, the parasite loses its specific invasion organelles (apical complex and surface coat) and de-differentiates into a round trophozoite located within a parasitophorous vacuole in the red blood cell cytoplasm. The young trophozoite (or "ring" stage, because of its morphology on stained blood films) grows substantially before undergoing schizogonic division.
During asexual development the parasite increases in size to ~50% of the uninfected erythrocyte volume: the infected erythrocyte volume remains relatively constant. Haemoglobin content gradually decreases but its concentration remains constant until the early trophozoite stage when it decreases by 25%. It then remains constant again until just prior to rupture. During early sexual development the gametocyte has a similar morphology to a trophozoite but subsequently undergoes a dramatic shape change.
Within the red blood cell, the parasite metabolism depends greatly on the digestion of hemoglobin
Hemoglobin
Hemoglobin is the iron-containing oxygen-transport metalloprotein in the red blood cells of all vertebrates, with the exception of the fish family Channichthyidae, as well as the tissues of some invertebrates...
. A set of enzymes known as plasmepsin
Plasmepsin
Plasmepsins are a class of at least 10 enzymes produced by the plasmodium parasite. There are ten different isoforms of these proteins and ten genes coding them respectively in plasmodium falciparum . It has been suggested that the plasmpesin family is smaller in other human plasmodium species...
s which are aspartic acid proteases
Protease
A protease is any enzyme that conducts proteolysis, that is, begins protein catabolism by hydrolysis of the peptide bonds that link amino acids together in the polypeptide chain forming the protein....
are used to degrade hemoglobin
Hemoglobin
Hemoglobin is the iron-containing oxygen-transport metalloprotein in the red blood cells of all vertebrates, with the exception of the fish family Channichthyidae, as well as the tissues of some invertebrates...
. The parasite digests 70-80% of the erythrocyte's haemoglobin but utilizes only ~15% in de novo protein synthesis. Intraerythrocytic Plasmodium falciparum utilizes only a fraction of the amino acids derived from the digestion of host cell cytosol for the biosynthesis of its proteins. The excess amino acids are exported from the infected erythrocyte by new transport pathways created by the parasite. The reason proposed for this apparently excessive digestion of haemoglobin is the colloid-osmotic hypothesis which suggests that the digestion of haemoglobin increases the osmotic pressure within the infected erythrocyte leading to its premature rupture and subsequent death of the parasite. To avoid this fate much of the haemoglobin is digested and exported from the erythrocyte. This hypothesis has been experimentally confirmed.
Infected erythrocytes are often sequestered in various human tissues or organs, such as the heart, liver and brain. This is caused by parasite derived cell surface proteins being present on the red blood cell membrane and it is these proteins that bind to receptors on human cells. Sequestration in the brain causes cerebral malaria, a very severe form of the disease, which increases the victim's likelihood of death.
The parasite can also alter the morphology of the red blood cell causing knobs on the erythrocyte membrane.
Erythrocyte invasion and growth leads to activation of several distinct anion channels and a non-selective Ca2+-permeable cation channel. The non-selective cation channel's activation allows entry of Ca2+ and Na+. Absence of the channels is incompatible with pathogen survial. Although the mechanism of activation of these channels is not know it is presumed to be due to oxidation stree generated by the parasite because similar or identical channels are activated by oxidation of non-infected erythrocytes. Ca2+ entry stimulates an intraerythrocytic scramblase
Scramblase
Scramblase is a protein responsible for the translocation of phospholipids between the two monolayers of a lipid bilayer of a cell membrane. In humans, phospholipid scramblases constitute a family of five homologous proteins that are named as hPLSCR1–hPLSCR5. Scramblases are members of the...
that facilitates bi-directional phospholipid
Phospholipid
Phospholipids are a class of lipids that are a major component of all cell membranes as they can form lipid bilayers. Most phospholipids contain a diglyceride, a phosphate group, and a simple organic molecule such as choline; one exception to this rule is sphingomyelin, which is derived from...
migration across the bilayer. This results in an alternation of the cell membrane's phosphatidylserine
Phosphatidylserine
Phosphatidylserine is a phospholipid component, usually kept on the inner-leaflet of cell membranes by an enzyme called flippase...
asymmetry. Exposure of phosphatidylserine at the outer surface of the cell membrane is followed by binding to phosphatidylserine receptors on macrophage
Macrophage
Macrophages are cells produced by the differentiation of monocytes in tissues. Human macrophages are about in diameter. Monocytes and macrophages are phagocytes. Macrophages function in both non-specific defense as well as help initiate specific defense mechanisms of vertebrate animals...
s and the subsequent phagocytosis of the affected erythrocyte. It appears that the parasite because of its growth requirements is in a race to complete its life cycle before the infected erythrocyte is phagocytosed.
Schizont
At the schizont stage, the parasite replicates its DNA multiple times without cellular segmentation. These schizonts then undergo cellular segmentation and differentiation to form roughly 16-18 merozoite cells in the erythrocyte. The merozoites burst from the red blood cell, and proceed to infect other erythrocytes. The parasite is in the bloodstream for roughly 60 seconds before it has entered another erythrocyte.
This infection cycle occurs in a highly synchronous fashion, with roughly all of the parasites throughout the blood in the same stage of development. This precise clocking mechanism has been shown to be dependent on the human host's own circadian rhythm
Circadian rhythm
A circadian rhythm, popularly referred to as body clock, is an endogenously driven , roughly 24-hour cycle in biochemical, physiological, or behavioural processes. Circadian rhythms have been widely observed in plants, animals, fungi and cyanobacteria...
. Specifically, human body temperature changes as a result of the circadian rhythm, seem to play a role in the development of P. falciparum
Plasmodium falciparum
Plasmodium falciparum is a protozoan parasite, one of the species of Plasmodium that cause malaria in humans. It is transmitted by the female Anopheles mosquito. Malaria caused by this species is the most dangerous form of malaria, with the highest rates of complications and mortality...
within the erythrocytic stage.
The synchronicity of the erythrocytic cycle is at least in part dependent on melatonin
Melatonin
Melatonin , also known chemically as N-acetyl-5-methoxytryptamine, is a naturally occurring compound found in animals, plants, and microbes...
secretion by the host. A mechanism for this has been proposed. Melatonin can activate phospholipase C which acts to generate inositol trisphosphate (IP3) which opens IP3 sensitive calcium
Calcium
Calcium is the chemical element with the symbol Ca and atomic number 20. It has an atomic mass of 40.078 amu. Calcium is a soft gray alkaline earth metal, and is the fifth-most-abundant element by mass in the Earth's crust...
channels in the endoplasmic reticulum
Endoplasmic reticulum
The endoplasmic reticulum is an organelle of cells in eukaryotic organisms that forms an interconnected network of tubules, vesicles, and cisternae...
. The released calcium in its turn controls the cycle through mechanisms that have yet to be understood.
Gametocyte differentiation
During the erythrocytic stage, some merozoites develop into male and female gametocytes. This process is called gametocytogenesis. The specific factors and causes underlying this sexual differentiation are largely unknown. These gametocytes take roughly 8–10 days to reach full maturity. Note that the gametocytes remain within the erythrocytes until taken up by the mosquitoMosquito
Mosquitoes are members of a family of nematocerid flies: the Culicidae . The word Mosquito is from the Spanish and Portuguese for little fly...
host.
Mosquito stage
P. falciparumPlasmodium falciparum
Plasmodium falciparum is a protozoan parasite, one of the species of Plasmodium that cause malaria in humans. It is transmitted by the female Anopheles mosquito. Malaria caused by this species is the most dangerous form of malaria, with the highest rates of complications and mortality...
is taken up by the female Anopheles
Anopheles
Anopheles is a genus of mosquito. There are approximately 460 recognized species: while over 100 can transmit human malaria, only 30–40 commonly transmit parasites of the genus Plasmodium, which cause malaria in humans in endemic areas...
mosquito
Mosquito
Mosquitoes are members of a family of nematocerid flies: the Culicidae . The word Mosquito is from the Spanish and Portuguese for little fly...
as it takes its bloodmeal from an infected human.
Gametogenesis
Upon being taken up by the mosquitoMosquito
Mosquitoes are members of a family of nematocerid flies: the Culicidae . The word Mosquito is from the Spanish and Portuguese for little fly...
, the gametocytes leave the erythrocyte shell and differentiate into gametes. The female gamete maturation process entails slight morphological changes, as it becomes enlarged and spherical. On the other hand, the male gamete maturation involves significant morphological development. The male gamete's DNA
DNA
Deoxyribonucleic acid is a nucleic acid that contains the genetic instructions used in the development and functioning of all known living organisms . The DNA segments that carry this genetic information are called genes, but other DNA sequences have structural purposes, or are involved in...
divides three times to form eight nuclei. Concurrently, eight flagella are formed. Each flagella pairs with a nucleus to form a microgamete, which separates from the parasite cell. This process is referred to as exflagellation.
Gametogenesis has been shown to be caused by: 1) a sudden drop in temperature upon leaving the human host, 2) a rise in pH within the mosquito
Mosquito
Mosquitoes are members of a family of nematocerid flies: the Culicidae . The word Mosquito is from the Spanish and Portuguese for little fly...
, and 3) xanthurenic acid
Xanthurenic acid
Xanthurenic acid is a chemical shown to induce gametogenesis of Plasmodium falciparum, the parasite that causes malaria. It is found in the gut of the Anopheles mosquito....
within the mosquito.
Fertilization
Fertilization of the female gamete by the male gamete occurs rapidly after gametogenesis. The fertilization event produces a zygoteZygote
A zygote , or zygocyte, is the initial cell formed when two gamete cells are joined by means of sexual reproduction. In multicellular organisms, it is the earliest developmental stage of the embryo...
. The zygote
Zygote
A zygote , or zygocyte, is the initial cell formed when two gamete cells are joined by means of sexual reproduction. In multicellular organisms, it is the earliest developmental stage of the embryo...
then develops into an ookinete. The zygote
Zygote
A zygote , or zygocyte, is the initial cell formed when two gamete cells are joined by means of sexual reproduction. In multicellular organisms, it is the earliest developmental stage of the embryo...
and ookinete are the only diploid stages of P. falciparum
Plasmodium falciparum
Plasmodium falciparum is a protozoan parasite, one of the species of Plasmodium that cause malaria in humans. It is transmitted by the female Anopheles mosquito. Malaria caused by this species is the most dangerous form of malaria, with the highest rates of complications and mortality...
.
Ookinete
The diploid ookinete is an invasive form of P. falciparumPlasmodium falciparum
Plasmodium falciparum is a protozoan parasite, one of the species of Plasmodium that cause malaria in humans. It is transmitted by the female Anopheles mosquito. Malaria caused by this species is the most dangerous form of malaria, with the highest rates of complications and mortality...
within the mosquito
Mosquito
Mosquitoes are members of a family of nematocerid flies: the Culicidae . The word Mosquito is from the Spanish and Portuguese for little fly...
. It traverses the peritrophic membrane of the mosquito
Mosquito
Mosquitoes are members of a family of nematocerid flies: the Culicidae . The word Mosquito is from the Spanish and Portuguese for little fly...
midgut and cross the midgut epithelium. Once through the epithelium, the ookinete enters the basal lamina, and forms an oocyst.
The processes of maturation and invasion of the mosquito gut are under investigation. Both chitinase
Chitinase
Chitinases are hydrolytic enzymes that break down glycosidic bonds in chitin. As chitin is a component of the cell walls of fungi and exoskeletal elements of some animals , chitinases are generally found in organisms that either need to reshape their own chitin or dissolve and digest the chitin of...
and plasmepsin
Plasmepsin
Plasmepsins are a class of at least 10 enzymes produced by the plasmodium parasite. There are ten different isoforms of these proteins and ten genes coding them respectively in plasmodium falciparum . It has been suggested that the plasmpesin family is smaller in other human plasmodium species...
4 (an aspartic acid protease
Protease
A protease is any enzyme that conducts proteolysis, that is, begins protein catabolism by hydrolysis of the peptide bonds that link amino acids together in the polypeptide chain forming the protein....
) are known to be involved in the invasion process.
During the ookinete stage, genetic recombination can occur. This takes place if the ookinete was formed from male and female gametes derived from different populations. This can occur if the human host contained multiple populations of the parasite, or if the mosquito
Mosquito
Mosquitoes are members of a family of nematocerid flies: the Culicidae . The word Mosquito is from the Spanish and Portuguese for little fly...
fed from multiple infected individuals within a short time-frame.
A G-actin binding protein has been implicated in this process. These proteins - probably acting as dimers - bind actin monomers just before sporogony.
Azithromycin
Azithromycin
Azithromycin is an azalide, a subclass of macrolide antibiotics. Azithromycin is one of the world's best-selling antibiotics...
has been shown to suppress apicoplast replication at the period of sporozoite production in oocysts.
The proteins enolase
Enolase
Enolase, also known as phosphopyruvate dehydratase, is a metalloenzyme responsible for the catalysis of the conversion of 2-phosphoglycerate to phosphoenolpyruvate , the ninth and penultimate step of glycolysis. Enolase belongs to the class Lyase. Enolase can also catalyze the reverse reaction,...
and actin
Actin
Actin is a globular, roughly 42-kDa moonlighting protein found in all eukaryotic cells where it may be present at concentrations of over 100 μM. It is also one of the most highly-conserved proteins, differing by no more than 20% in species as diverse as algae and humans...
are present on the surface of ookinetes but their function there, if any, is unknown.
The myosin gene (MyoA) class XIV is essential for ookinete motility in the mosquito. Mutants placed under a different promotor active in the blood stages failed to complete their life cycle. Disruption serine repeat antigen 5 blocks parasite inhibits egress of sporozoites from an oocyst.
Sporogony
Over the period of a 1–3 weeks, the oocyst grows to a size of tens to hundreds of micrometres. During this time, multiple nuclear divisions occur. After oocyst maturation is complete, the oocyst divides to form multiple haploid sporozoites. Immature sporozoites break through the oocyst wall into the haemolymph. The sporozoites then migrate to the salivary glands and complete their differentiation. Once mature, the sporozoites can proceed to infect a human host during a subsequent mosquitoMosquito
Mosquitoes are members of a family of nematocerid flies: the Culicidae . The word Mosquito is from the Spanish and Portuguese for little fly...
bite.
Morphology
The nucleus, mitochondrion, apicoplast and the microtubuleMicrotubule
Microtubules are a component of the cytoskeleton. These rope-like polymers of tubulin can grow as long as 25 micrometers and are highly dynamic. The outer diameter of microtubule is about 25 nm. Microtubules are important for maintaining cell structure, providing platforms for intracellular...
s of Plasmodium sporozoites are linked to the parasite pellicle
Pellicle (biology)
The pellicle is a thin layer supporting the cell membrane in various protozoa, protecting them and allowing them to retain their shape, especially during locomotion, allowing the organism to be more hydrodynamic. They vary from flexible and elastic to rigid. Although stiff, the pellicle is...
via long tethering proteins. The tethers originate from the inner membrane complex and are arranged in a periodic fashion following a 32 nanometer repeat. The tethers pass through a subpellicular structure that encompasses the entire parasite, probably as a network of membrane associated filaments.
Cell division
Cell division occurs through a process known as schizogony. This is a type of mitotic division in which multiple rounds of nuclear divisions occur before the cytoplasm segments.DNA synthesis begins in the relatively small trophozoites but nuclear subdivision, which leads to the formation of multinucleate cells, occurs only during schizogony. Whether or not any gap phases exist between each round of DNA synthesis and mitosis is unknown. Eventually, a schizont composed of 8–32 nuclei undergoes segmentation, which culminates with the formation of individual merozoites that burst from the erythrocyte into the blood stream.
Liver stages
Invasion of the hepatocytes appears to involve at least 2 proteins: sporozoite invasion-associated proteins (SIAP)-1 and -2. These proteins bind heparinHeparin
Heparin , also known as unfractionated heparin, a highly sulfated glycosaminoglycan, is widely used as an injectable anticoagulant, and has the highest negative charge density of any known biological molecule...
sulfate and chondroitin
Chondroitin
Chondroitin is a chondrin derivative.Types include:* Chondroitin sulfate* Dermatan sulfate...
sulfate type membrane receptors on host cells.
Latency of sporozoites is controlled by the eIF2
EIF2
Eukaryotic Initiation Factor 2 is a eukaryotic initiation factor. It is required in the initiation of translation. In this fundamental process of life, the ribosome builds proteins according to the information encoded on the mRNA. eIF2 mediates the binding of tRNAmet to the ribosome in a...
alpha kinase IK2, a general inhibitor of protein synthesis. Puf2 participates in the regulation of IK2 and inhibits premature sporozoite transformation. In contrast Puf1 appears to be dispensable.
The RNA binding protein family PUF member Pumilio-2 (Puf2) appears to be involved in transformation of sporozoites into the hepatic stage of the life cycle. Knock out mutants of this gene exhibit genome wide transcriptional changes resulting in loss of gliding motility, cell traversal ability, reduction in infectivity and trigger metamorphosis typical of early Plasmodium intra-hepatic development.
The division of the liver stages into thousands of merozoites is a complex process. In parallel with nuclear division, the apicoplast and mitochondrion become two extensively branched and intertwining structures. The organelles subsequently undergo morphological and positional changes prior to cell division. Finally to form merozoites, the parasite undergoes cytokinesis.
Type II fatty acid
Fatty acid
In chemistry, especially biochemistry, a fatty acid is a carboxylic acid with a long unbranched aliphatic tail , which is either saturated or unsaturated. Most naturally occurring fatty acids have a chain of an even number of carbon atoms, from 4 to 28. Fatty acids are usually derived from...
biosynthesis is vital for this stage in the life cycle. This pathway may be inhibited by the antibiotic triclosan
Triclosan
Triclosan is an antibacterial and antifungal agent. It is a polychloro phenoxy phenol. Despite being used in many consumer products, beyond its use in toothpaste to prevent gingivitis, there is no evidence according to the American Food and Drug Administration that triclosan provides an extra...
.
During this stage of development the sporozoite selectively discards organelles unnecessary for growth at this stage of the life cycle. Among these are the micronemes and the inner membrane complex.
The host iron regulatory hormone hepcidin
Hepcidin
Hepcidin is a peptide hormone produced by the liver. It was discovered in 2000, and appears to be the master regulator of iron homeostasis in humans and other mammals. In humans, HAMP is the gene that encodes for hepcidin.-Structure:...
which is synthesised in the liver and spleen
Spleen
The spleen is an organ found in virtually all vertebrate animals with important roles in regard to red blood cells and the immune system. In humans, it is located in the left upper quadrant of the abdomen. It removes old red blood cells and holds a reserve of blood in case of hemorrhagic shock...
, appears to be able to inhibit growth of the liver stages. Levels of this hormone are elevated during infection and seem to correlate with the anaemia often found in malaria. Erythrocytic parasitaemia, above a minimum threshold, impairs the growth of subsequent liver cell sporozoite infection. The production of hepcidin leads to the redistributes iron away from hepatocytes thus slowing the development of the iron dependent liver stage.
Liver hepcidin expression is upregulated and downregulated during the early and late stages of malaria infection respectively. Inflammation and erythropoietin, rather than the iron sensing pathway, are involved in the regulation of hepcidin expression. Treatment of malaria infected mice with anti hepcidin neutralizing antibodies increased parasitemia and mortality rates. Overexpression of hepcidin improves the outcome.
Erythrocyte invasion
This is a complex and poorly understood process. The merozoite initially contacts the erythrocyte and rotates until the rhoptery containing part is adjacent to the erythrocyte membrane. A tight contact is then established and the parasite enters the erythrocyte. This happens within seconds making the invasion process difficult to analyse.Two families of proteins are known to be involved in this process: the reticulocyte binding-like homologues (PfRh or PfRBP) and erythrocyte binding-like (EBL) proteins. The PfRh family consists of five proteins and a pseudogene
Pseudogene
Pseudogenes are dysfunctional relatives of known genes that have lost their protein-coding ability or are otherwise no longer expressed in the cell...
: PfRh1, PfRh2a, PfRh2b, PfRh3, PfRh4 and PfRh5. PfRh3 is a transcribed pseudogene in all strains examined to date. All the other members of this family bind to erythrocyes and antibodies to them inhibit invasion. PfRh5 is located within the rhoptries and appears to be an essential gene.
The EBL family of proteins includes EBA-175, EBA-181 (also known as JESEBL), EBA-140 (also known as BAEBL) and EBL-1. Whilst these parasite ligands function in merozoite invasion by binding to specific receptors on the erythrocyte, they appear also to have a central role in activation of the invasion process. Binding of EBA-175 to its receptor, glycophorin A
GYPA
Glycophorin A , also known as GYPA, is a protein which in humans is encoded by the GYPA gene. GYPA has also recently been designated CD235a .- Function :...
, restores the basal cytosolic calcium levels after interaction of the merozoite with the erythrocyte and triggers the release of rhoptry proteins.
There seems to be some overlap between the functions of these proteins. Loss of EBA-175 can be compensated by increased expression of PfRh4.
Some details of the invasion process are known. The rhoptery protein RON2 is inserted into the erythrocyte membrane. The protein AMA1 secreted from microneme then binds to RON2. RON2 forms part of a macromolecular complex which includes RON2, RON 4, RON5 and RON8.
The residues of the RON2 protein binds to the AMA-1 protein have been identified. It also appears that the formation of the junction and parasitophorous vacuole are molecularly distinct steps in the invasion process.
The invasion process appears to be ATP dependent and may involve a purogenic signalling pathway.
Reticulocyte binding like protein homologue 2a (PfRH2a) is processed both in the schizont as well as during invasion resulting in proteins with different erythrocyte binding properties. It also moves from the rhoptry neck to the moving junction during merozoite invasion. PfRh2a undergoes a cleavage event in the transmembrane region during invasion consistent with activity of the membrane associated PfROM4 protease. Both PfRh2a and PfRh2b bind to red blood cells. The erythrocyte-binding domain lies within a 15 kDa region at the N-terminus of each protein.
A GPI-anchored micronemal antigen (GAMA) appears to be essential in the process of erythrocyte invasion.
PfRh4 binds to a second protein P. falciparum Rh5 interacting protein (PfRipr). PfRipr has a molecular weight of 123 kiloDaltons with 10 epidermal growth factor
Epidermal growth factor
Epidermal growth factor or EGF is a growth factor that plays an important role in the regulation of cell growth, proliferation, and differentiation by binding to its receptor EGFR...
-like domains and 87 cysteine
Cysteine
Cysteine is an α-amino acid with the chemical formula HO2CCHCH2SH. It is a non-essential amino acid, which means that it is biosynthesized in humans. Its codons are UGU and UGC. The side chain on cysteine is thiol, which is polar and thus cysteine is usually classified as a hydrophilic amino acid...
residues distributed along the protein.
The P. falciparum apical sushi protein is the homolog of the P. vivax RON1 protein.
The receptor for the PfRh5 protein appears to be the Ok blood group antigen, basigin
Basigin
Basigin also known as extracellular matrix metalloproteinase inducer or cluster of differentiation 147 is a protein that in humans is encoded by the BSG gene. This protein is a determinant for the Ok blood group system...
. Blocking access to this protein on the erythrocyte surface appears to inhibit erythrocyte invasion completely. Binding of the Rh5 protein appears to be critically dependent on a single residue within the Rh5 protein.
Apical complex
Several proteins are involved in the binding of the sporozoite to the various tissues it attaches to. TRAPAttenuator (genetics)
Attenuation is a regulatory feature found throughout Archaea and Bacteria causing premature termination of transcription. Attenuators are 5'-cis acting regulatory regions which fold into one of two alternative RNA structures which determine the success of transcription...
, S6 and TLP have been implicated in these processes. Heparin
Heparin
Heparin , also known as unfractionated heparin, a highly sulfated glycosaminoglycan, is widely used as an injectable anticoagulant, and has the highest negative charge density of any known biological molecule...
like molecules bound to the surface of the erythrocyte appear to be important in this process which involves the merozoite surface protein 1.
The rhoptery neck proteins (RONs) along with the micronemal AMA1 protein are important in the penetration of the erythrocyte. These form part of the moving junction which initially binds to the erythrocyte surface and is involved in the entry of the parasite into the erythrocyte cytoplasm. The mechanisms involved in this process are still being elucidated. The protein RON8 appears to be central to the binding of parasite to the erythrocyte surface.
Maurer's clefts
In 1902 the German physician Georg Maurer discovered an unusual staining pattern in the cytoplasm of erythrocytes infected with P. falciparum. These structures were subsequently named Maurer's clefts. These consist of a convoluted set of membranes that lie within the erythrocte's cytoplasm and appear to be involved in secrection from the erythrocyte. They are known to have proteins of parasite origin within them including the Maurer's cleft two transmembrane proteins (PfMC-2TM) The clefts appear to originate from vacuoles budding off them the parasitophorous vacuole membrane which then diffuse within the erythrocyte cytoplasm before taking up residence at the cell periphery.Another protein associated with these structure is skeleton-binding protein 1 (SBP1). This protein is invovled in transport of the var gene protein (erythrocyte membrane protein 1) to the erythrocyte surface.
Mutations in the ring exported protein 1 (Rex 1), a protein normally found in Maurer's clefts, reduces transport ofthe var gene products to the erythrocyte surface.
Parasitophorous vacuole
Within a red blood cell, P. falciparumPlasmodium falciparum
Plasmodium falciparum is a protozoan parasite, one of the species of Plasmodium that cause malaria in humans. It is transmitted by the female Anopheles mosquito. Malaria caused by this species is the most dangerous form of malaria, with the highest rates of complications and mortality...
resides inside the parasitophorous vacuole. This is formed during erythrocyte invasion.
The proteins originating in the parasite pass through the membrane of the parasitophorous vacuole and are transported to the cytoplasm or membrane of the erythrocyte. Although this transport mechanism is largely unknown some details have been elucidated. Ingestion of the erythrocyte cytoplasm begins in mid-ring-stage parasites. Host cytoplasm
Cytoplasm
The cytoplasm is a small gel-like substance residing between the cell membrane holding all the cell's internal sub-structures , except for the nucleus. All the contents of the cells of prokaryote organisms are contained within the cytoplasm...
is internalised via cytostome
Cytostome
A cytostome or cell mouth is a part of a cell specialized for phagocytosis, usually in the form of a microtubule-supported funnel or groove. Food is directed into the cytostome, and sealed into vacuoles. Only certain groups of protozoa, such as the ciliates and excavates, have cytostomes. Such...
-derived invaginations and then concentrated into several acidified peripheral structures. Haemoglobin digestion and haemozoin
Hemozoin
Hemozoin is a disposal product formed from the digestion of blood by some blood-feeding parasites. These hematophagous organisms such as Malaria parasites , Rhodnius and Schistosoma digest hemoglobin and release high quantities of free heme, which is the non-protein component of hemoglobin...
formation occur within these vesicle
Vesicle (biology)
A vesicle is a bubble of liquid within another liquid, a supramolecular assembly made up of many different molecules. More technically, a vesicle is a small membrane-enclosed sack that can store or transport substances. Vesicles can form naturally because of the properties of lipid membranes , or...
s. The ring-stage parasites can adopt a deeply invaginated cup shape, but they do not take up haemoglobin via macropinocytosis. As the parasite matures the haemozoin containing compartments coalesce to form a single acidic digestive vacuole (pH 4.5 - 5.5) that is then fed by haemoglobin containing vesicles. Some haemoglobin degradation also occurs in compartments outside the digestive vacuole.
The enzyme phosphatidyl-inositol-3-kinase (PI3K) has been implicated in this process. PI3K is located in vesicular compartments near the membrane and in the food vacuole and is involved in endocytosis from the host and trafficking of hemoglobin in the parasite. Its inhibition with wortmannin
Wortmannin
Wortmannin, a furanosteroid metabolite of the fungi Penicillium funiculosum, Talaromyces wortmannii,is a specific, covalent inhibitor of phosphoinositide 3-kinases . It has an in vitro inhibitory concentration of around 5 nM, making it a more potent inhibitor than LY294002, another commonly...
or LY294002
LY294002
LY294002 is a morpholine derivative of quercetin. It is a potent inhibitor of phosphoinositide 3-kinases . Two of these are the proto-oncogene serine/threonine-protein kinase and the phosphatidylinositol-4,5-bisphosphate 3-kinase catalytic subunit gamma isoform...
results in entrapment of hemoglobin in vesicles within the parasite cytoplasm preventing its transport to the food vacuole.
The pH of the digestive vacuole is maintained by a V-type H(+)-ATPase.
A signal sequence at the N terminal of proteins targeted to the arasitophorous vacuole has been identified. The signal appears to reside in the 55 amino acids of the N terminal of the protein. There may be a retention signal at the C terminal.
The micronemal protease ROM1 appears to be essential for proper parasitophorous vacuole modification to allow parasite development. This protease is able to cleave the proteins AMA1 and MAEBL.
Apicoplast
Plasmodium falciparumPlasmodium falciparum
Plasmodium falciparum is a protozoan parasite, one of the species of Plasmodium that cause malaria in humans. It is transmitted by the female Anopheles mosquito. Malaria caused by this species is the most dangerous form of malaria, with the highest rates of complications and mortality...
, and most other members of the phylum
Phylum
In biology, a phylum The term was coined by Georges Cuvier from Greek φῦλον phylon, "race, stock," related to φυλή phyle, "tribe, clan." is a taxonomic rank below kingdom and above class. "Phylum" is equivalent to the botanical term division....
Apicomplexa
Apicomplexa
The Apicomplexa are a large group of protists, most of which possess a unique organelle called apicoplast and an apical complex structure involved in penetrating a host's cell. They are unicellular, spore-forming, and exclusively parasites of animals. Motile structures such as flagella or...
, contain an organelle
Organelle
In cell biology, an organelle is a specialized subunit within a cell that has a specific function, and is usually separately enclosed within its own lipid bilayer....
termed an apicoplast
Apicoplast
An apicoplast is a derived non-photosynthetic plastid found in most Apicomplexa, including malaria parasites such as Plasmodium falciparum, but not in others such as Cryptosporidium. It originated from an algae through secondary endosymbiosis...
. The apicoplast
Apicoplast
An apicoplast is a derived non-photosynthetic plastid found in most Apicomplexa, including malaria parasites such as Plasmodium falciparum, but not in others such as Cryptosporidium. It originated from an algae through secondary endosymbiosis...
is an essential
Essential (biology)
An essential part of an organism is something that the organism cannot continue to be alive or reproduce without. For instance, mitochondria are essential to most eukaryotic cells. Genes can also be considered with regards to their essentiality...
plastid
Plastid
Plastids are major organelles found in the cells of plants and algae. Plastids are the site of manufacture and storage of important chemical compounds used by the cell...
, homologous to a chloroplast
Chloroplast
Chloroplasts are organelles found in plant cells and other eukaryotic organisms that conduct photosynthesis. Chloroplasts capture light energy to conserve free energy in the form of ATP and reduce NADP to NADPH through a complex set of processes called photosynthesis.Chloroplasts are green...
, although the apicoplast
Apicoplast
An apicoplast is a derived non-photosynthetic plastid found in most Apicomplexa, including malaria parasites such as Plasmodium falciparum, but not in others such as Cryptosporidium. It originated from an algae through secondary endosymbiosis...
is not photosynthetic. Evolutionarily, it is thought to have derived through secondary endosymbiosis.
The function of the apicoplast
Apicoplast
An apicoplast is a derived non-photosynthetic plastid found in most Apicomplexa, including malaria parasites such as Plasmodium falciparum, but not in others such as Cryptosporidium. It originated from an algae through secondary endosymbiosis...
remains to be fully determined, but it appears to be involved in the metabolism of fatty acids, isoprenoids, and heme
Heme
A heme or haem is a prosthetic group that consists of an iron atom contained in the center of a large heterocyclic organic ring called a porphyrin. Not all porphyrins contain iron, but a substantial fraction of porphyrin-containing metalloproteins have heme as their prosthetic group; these are...
.
The apicoplast
Apicoplast
An apicoplast is a derived non-photosynthetic plastid found in most Apicomplexa, including malaria parasites such as Plasmodium falciparum, but not in others such as Cryptosporidium. It originated from an algae through secondary endosymbiosis...
contains a 35-kb genome
Genome
In modern molecular biology and genetics, the genome is the entirety of an organism's hereditary information. It is encoded either in DNA or, for many types of virus, in RNA. The genome includes both the genes and the non-coding sequences of the DNA/RNA....
, which encodes for 30 proteins. Other, nuclear-encoded, proteins are transported into the apicoplast
Apicoplast
An apicoplast is a derived non-photosynthetic plastid found in most Apicomplexa, including malaria parasites such as Plasmodium falciparum, but not in others such as Cryptosporidium. It originated from an algae through secondary endosymbiosis...
using a specific signal peptide
Signal peptide
A signal peptide is a short peptide chain that directs the transport of a protein.Signal peptides may also be called targeting signals, signal sequences, transit peptides, or localization signals....
. It is estimated that 551, or roughly 10%, of the predicted nuclear-encoded proteins are targeted to the apicoplast
Apicoplast
An apicoplast is a derived non-photosynthetic plastid found in most Apicomplexa, including malaria parasites such as Plasmodium falciparum, but not in others such as Cryptosporidium. It originated from an algae through secondary endosymbiosis...
.
As humans do not harbor apicoplasts, this organelle and its constituents are seen as a possible target for antimalarial drugs.
The plastid genome replicates at the late trophozoite stage of the parasite intraerythocytic cycle. It proceeds predominantly via a D-loop/bi-directional ori mechanism with replication ori localized within the inverted repeat region. The process of replication involves a nuclear-encoded DnaJ homolog that binds to the ori site.
The DNA polymerase
DNA polymerase
A DNA polymerase is an enzyme that helps catalyze in the polymerization of deoxyribonucleotides into a DNA strand. DNA polymerases are best known for their feedback role in DNA replication, in which the polymerase "reads" an intact DNA strand as a template and uses it to synthesize the new strand....
involved in the replication of its genome is Pfprex (Klenow-like polymerase). This enzyme has been cloned, expressed and purified. The enzymes is relatively error prone and shows a bias toward T->C mutauons.
The bisynthesis of this organelle is not well understood. Phosphatidylinositol 3-monophosphate has been shown to be involved in its biosynthesis in the apicomplexian Toxoplasma gondii
Toxoplasma gondii
Toxoplasma gondii is a species of parasitic protozoa in the genus Toxoplasma. The definitive host of T. gondii is the cat, but the parasite can be carried by many warm-blooded animals . Toxoplasmosis, the disease of which T...
. It seems likely that this enzyme is involved in the formation of this organells in the Plasmodium species also.
Transport into the apicoplast are not well understood. These proteins has a signal in the N terminal but unlike many other organisms this appears to be a disordered chain rather than a conserved sequence.
Iron-sulphur prosthetic groups are assembled in this organelle. One component (SufB) is encoded in the apicoplast genome and a second (SufC
SUFC
SUFC can refer to one of the following:* Scunthorpe United F.C.* Seoul United FC* Sheffield United F.C.* Southend United F.C.* Spalding United F.C.* Supersport United F.C.* Sutton United F.C....
) is encoded in the nucleus. SufB also exhibits ATPase activity. Other pathways that have been linked to this organelle include biosynthesis of isoprenoid precursors, fatty acids, heme and lipoic acid.
A gene Plasmodium-specific Apicoplast protein for Liver Merozoite formation (PALM) has been shown to be important for merozoite formation. Knock out mutants are unable to release merozoites into the blood from the liver stages. Mutants lacking this gene appear to be able to elicit at least temporary immunity.
The role of the apicoplast in the blood stages has been clarified. Inhibition of isoprenoid precursor biosynthesis with the antibiotic fosmidomycin
Fosmidomycin
Fosmidomycin is an antibiotic that was originally isolated from culture broths of bacteria of the genus Streptomyces. It specifically inhibits DXP reductoisomerase, a key enzyme in the non-mevalonate pathway of isoprenoid biosynthesis. It is a structural analogue of 2-C-methyl-D-erythrose...
(an inhibitor of the enzyme DOXP reductoisomerase) causes delayed death in this parasite. This effect can be overcome with the addition of isopentenyl pyrophosphate
Isopentenyl pyrophosphate
Isopentenyl pyrophosphate is an intermediate in the classical, HMG-CoA reductase pathway used by organisms in the biosynthesis of terpenes and terpenoids. IPP is formed from acetyl-CoA via mevalonic acid...
(IPP) to the culture medium. Continued culture in the presence of this agent leads to the loss of the apicoplast genome and these mutants fail to process or localize organelle proteins. These auxotrophs can be grown indefinitely in asexual blood stage culture but are entirely dependent on exogenous IPP for survival.
Mitochondrion
Plasmodium lacks mitochondrial pyruvate dehydrogenasePyruvate dehydrogenase
Pyruvate dehydrogenase complex is a complex of three enzymes that transform pyruvate into acetyl-CoA by a process called pyruvate decarboxylation. Acetyl-CoA may then be used in the citric acid cycle to carry out cellular respiration, and this complex links the glycolysis metabolic pathway to the...
and the hydrogen ion translocating NADH dehydrogenase
Dehydrogenase
A dehydrogenase is an enzyme that oxidises a substrate by a reduction reaction that transfers one or more hydrides to an electron acceptor, usually NAD+/NADP+ or a flavin coenzyme such as FAD or FMN.-Examples:...
(Complex I, NDH1). The mitochondrion
Mitochondrion
In cell biology, a mitochondrion is a membrane-enclosed organelle found in most eukaryotic cells. These organelles range from 0.5 to 1.0 micrometers in diameter...
contains a minimal DNA genome (~6 kilobases) and carries out oxidative phosphorylation in the insect vector stages by using 2-oxoglutarate as an alternative means of entry into the tricarboxylic acid cycle and a single-subunit flavoprotein as an alternative NADH dehydrogenase (NDH2). In the blood stages mitochondrial enzymes are down regulated and parasite energy metabolism relies mainly on glycolysis.
The ATP synthase
ATP synthase
right|thumb|300px|Molecular model of ATP synthase by X-ray diffraction methodATP synthase is an important enzyme that provides energy for the cell to use through the synthesis of adenosine triphosphate . ATP is the most commonly used "energy currency" of cells from most organisms...
is localised to the mitochondrion, is assembled as a large dimeric complex and appears to be essential for in the blood stages of the life cycle. Its function in these stages is not yet clear.
The dicarboxylate-tricarboxylate carrier homolog has been cloned from P. falciparum. This protein may mediate the oxoglutarate-malate
Malate
Malate is the ionized form of malic acid. It is an important chemical compound in biochemistry. In the C4 carbon fixation process, malate is a source of CO2 in the Calvin cycle....
exchange across the inner mitochondrial membrane required for the branched pathway of tricarboxylic acid metabolism.
The ClpQ protease and ClpY ATPase have been cloned. ClpQY function disruption caused hindrance in the parasite growth and maturation of asexual stages of parasites. Features of apoptosis like cell death are also found.
Digestive vacuole
It has been shown that micromolar concentrations of chloroquineChloroquine
Chloroquine is a 4-aminoquinoline drug used in the treatment or prevention of malaria.-History:Chloroquine , N'--N,N-diethyl-pentane-1,4-diamine, was discovered in 1934 by Hans Andersag and co-workers at the Bayer laboratories who named it "Resochin". It was ignored for a decade because it was...
partially permeabilized the parasite's digestive vacuole membrane and that this event appears to precede mitochondrial dysfunction.
Nucleolus
A hat-like structure polarized towards one side of the nucleus that stains with nucleolar markers has been described It seems likely that this unusual structure is the nucleolusNucleolus
The nucleolus is a non-membrane bound structure composed of proteins and nucleic acids found within the nucleus. Ribosomal RNA is transcribed and assembled within the nucleolus...
.
Ribosomes
Unlike other eukaryotes studied to date PlasmodiumPlasmodium
Plasmodium is a genus of parasitic protists. Infection by these organisms is known as malaria. The genus Plasmodium was described in 1885 by Ettore Marchiafava and Angelo Celli. Currently over 200 species of this genus are recognized and new species continue to be described.Of the over 200 known...
species have two or three distinct SSU rRNA (18S rRNA) molecules encoded within the genome. These have been divided into types A, S and O. Type A is expressed in the asexual stages; type S in the sexual and type O only in the oocyte. Type O is only known to occur in Plasmodium vivax
Plasmodium vivax
Plasmodium vivax is a protozoal parasite and a human pathogen. The most frequent and widely distributed cause of recurring malaria, P. vivax is one of the four species of malarial parasite that commonly infect humans. It is less virulent than Plasmodium falciparum, which is the deadliest of the...
at present. The reason for this gene duplication is not known but presumably reflects an adaption to the different environments the parasite lives within.
The Asian simian Plasmodium species - Plasmodium coatneyi, Plasmodium cynomolgi, Plasmodium fragile, Plasmodium inui
Plasmodium inui
Plasmodium inui is a protozoan parasite, one of the species of simian Plasmodium that cause malaria in Old World monkeys.-Hosts:* Assamese macques * long tailed macques -Life cycle:...
, Plasmodium fieldi
Plasmodium fieldi
Plasmodium fieldi is a parasite of the genus Plasmodium sub genus Plasmodium found in Malaysia.This species is related to Plasmodium ovale and Plasmodium simiovale. Like all Plasmodium species P. fieldi has both vertebrate and insect hosts...
, Plasmodium hylobati and Plasmodium simiovale - have a single single S-type-like gene and several A-type-like genes. Phylogenetic analyses has shown that gene duplication events giving rise to A- and S-type-like sequences took place independently at least three times in the Plasmodium evolution.
Sexual differentiation
Sexual parasite development is controlled by a DEAD boxDEAD box
DEAD box proteins are involved in an assortment of metabolic processes that involve RNA. They are highly conserved in nine domains and can be found in both prokaryotes and eukaryotes, but not all...
RNA
RNA
Ribonucleic acid , or RNA, is one of the three major macromolecules that are essential for all known forms of life....
helicase
Helicase
Helicases are a class of enzymes vital to all living organisms. They are motor proteins that move directionally along a nucleic acid phosphodiester backbone, separating two annealed nucleic acid strands using energy derived from ATP hydrolysis.-Function:Many cellular processes Helicases are a...
of the DDX6 family, termed DOZI.
The Puf2 gene, a member of the Puf family of transcriptional regulators, has been shown to be involved in gamete formation.
FACT
FACT (biology)
FACT is a heterodimeric protein complex that affects eukaryotic RNA polymerase II transcription elongation both in vitro and in vivo...
(facilitates chromatin transcription) is a dimeric complex of two protein
Protein
Proteins are biochemical compounds consisting of one or more polypeptides typically folded into a globular or fibrous form, facilitating a biological function. A polypeptide is a single linear polymer chain of amino acids bonded together by peptide bonds between the carboxyl and amino groups of...
s - SPT16
SUPT16H
FACT complex subunit SPT16 is a protein that in humans is encoded by the SUPT16H gene.-Further reading:...
and SSRP1 - which acts as a histone
Histone
In biology, histones are highly alkaline proteins found in eukaryotic cell nuclei that package and order the DNA into structural units called nucleosomes. They are the chief protein components of chromatin, acting as spools around which DNA winds, and play a role in gene regulation...
chaperone in the (dis)assembly of nucleosome
Nucleosome
Nucleosomes are the basic unit of DNA packaging in eukaryotes, consisting of a segment of DNA wound around a histone protein core. This structure is often compared to thread wrapped around a spool....
(and chromatin
Chromatin
Chromatin is the combination of DNA and proteins that make up the contents of the nucleus of a cell. The primary functions of chromatin are; to package DNA into a smaller volume to fit in the cell, to strengthen the DNA to allow mitosis and meiosis and prevent DNA damage, and to control gene...
) structure during transcription
Transcription (genetics)
Transcription is the process of creating a complementary RNA copy of a sequence of DNA. Both RNA and DNA are nucleic acids, which use base pairs of nucleotides as a complementary language that can be converted back and forth from DNA to RNA by the action of the correct enzymes...
and DNA replication. It is an essential gene in Plasmodium. Changing its promoter to one expressed only in the blood stages leads to changes in the male gametocytes. The mutant gametocytes have delayed DNA replication and gametocyte formation. Male gamete fertility is strongly reduced. Female gametocytes appear to be normal. When successful fertilization is achieved, the ookinetes generate oocysts that arrest early in development and fail to enter sporogony.
Egress from the erythrocyte
This is an essential step in the life cycle. The calcium dependent protein kinase PfCDPK5 which is expressed in the merozoite is essential for this process. Deletion mutations of this gene result cell arrest in the late schizont stages. Merozoites released from these schizonts are capable of invasion.Large holes appear in the cytoskeleton ~35 hours post invasion. This occurs at the same time as the loss of cytoskeletal adaptor proteins that are part of the junctional complex, including α/β-adducin and tropomyosin
Tropomyosin
Tropomyosin is an actin-binding protein that regulates actin mechanics. It is important, among other things, for muscle contraction. Tropomyosin, along with the troponin complex, associate with actin in muscle fibers and regulate muscle contraction by regulating the binding of myosin...
. This is followed by the proteolysis of many cytoskeletal proteins during egress at ~48 hours post infection. This later proteolysis is mediated by the erythrocyte's own calpain
Calpain
A calpain is a protein belonging to the family of calcium-dependent, non-lysosomal cysteine proteases expressed ubiquitously in mammals and many other organisms. Calpains constitute the C2 family of protease clan CA in the MEROPS database...
-1.
Along with the release from the erythrocyte of the merozoites, the now functionless digestive vacuole is also released. These are can active complement and are rapidly taken up by the polymorph
Granulocyte
Granulocytes are a category of white blood cells characterized by the presence of granules in their cytoplasm. They are also called polymorphonuclear leukocytes because of the varying shapes of the nucleus, which is usually lobed into three segments...
s. On ingestion the digestive vacuoles induce a vigorous respiratory burst which drives the cells into a state of functional exhaustion, blunting production of reactive oxygen species and microbicidal activity upon challenge with bacterial pathogens.
The serine repeat antigen (SERA) multigene family encode a series of proteins with a putative papain
Papain
Papain, also known as papaya proteinase I, is a cysteine protease enzyme present in papaya and mountain papaya .-Papain family:...
-like cysteine
Cysteine
Cysteine is an α-amino acid with the chemical formula HO2CCHCH2SH. It is a non-essential amino acid, which means that it is biosynthesized in humans. Its codons are UGU and UGC. The side chain on cysteine is thiol, which is polar and thus cysteine is usually classified as a hydrophilic amino acid...
protease
Protease
A protease is any enzyme that conducts proteolysis, that is, begins protein catabolism by hydrolysis of the peptide bonds that link amino acids together in the polypeptide chain forming the protein....
motif. One of these SERA5 (120 kiloDaltons) is produced at the late trophozoite/schizont stage. It is secreted together with other SERAs into the parasitophorous vacuole in an infected erythrocyte where it is cleaved into three fragments: an N-terminal domain (47 kDa), a central domain containing putative papain-like cysteine protease motifs (56 kDa) and a C-terminal domain (18 kDa). This N-terminal fragment is then cleaved in turn into two 25 kDa fragments. These fragments become covalently linked to the C-terminal 18 kDa fragment via disulfide bonding and attach to the merozoite surface. The central fragment is further cleaved to 50 kDa and 6 kDa fragments before being shed to the medium. These proteolytic cleavages are carried out by a subtilisin
Subtilisin
Subtilisin is a non-specific protease initially obtained from Bacillus subtilis.Subtilisins belong to subtilases, a group of serine proteases that initiate the nucleophilic attack on the peptide bond through a serine residue at the active site. They are physically and chemically...
-like serine
Serine
Serine is an amino acid with the formula HO2CCHCH2OH. It is one of the proteinogenic amino acids. By virtue of the hydroxyl group, serine is classified as a polar amino acid.-Occurrence and biosynthesis:...
protease called PfSUB1 and the inhibition of this processing, likewise, results in blockade of merozoite release. SERA6 may also be involved in schizont rupture and merozoite release from the erythrocyte.
A protein - gamete egress and sporozoite traversal - has been identified that appears to be involved in the egress of male and female gametes from the erythrocyte. It is also involved in sporozoite migration.
Kinases
Although several kinaseKinase
In chemistry and biochemistry, a kinase is a type of enzyme that transfers phosphate groups from high-energy donor molecules, such as ATP, to specific substrates, a process referred to as phosphorylation. Kinases are part of the larger family of phosphotransferases...
s are known in P. falciparum very little is known about them. Several are cyclin
Cyclin
Cyclins are a family of proteins that control the progression of cells through the cell cycle by activating cyclin-dependent kinase enzymes.- Function :...
dependent kinase kinase like kinases (CLK): of these two - the Lammer kinase homologue PfCLK-1 and PfCLK-2 have been cloned. CLKs in other eukaryotes are involved in the regulation of mRNA splicing through phosphorylation of serine
Serine
Serine is an amino acid with the formula HO2CCHCH2OH. It is one of the proteinogenic amino acids. By virtue of the hydroxyl group, serine is classified as a polar amino acid.-Occurrence and biosynthesis:...
/arginine
Arginine
Arginine is an α-amino acid. The L-form is one of the 20 most common natural amino acids. At the level of molecular genetics, in the structure of the messenger ribonucleic acid mRNA, CGU, CGC, CGA, CGG, AGA, and AGG, are the triplets of nucleotide bases or codons that codify for arginine during...
-rich proteins. Both are transcribed throughout the asexual blood stages and in gametocytes. PfCLK-1/Lammer possesses two nuclear localization signal sites while PfCLK-2 possesses one of these signal sites upstream of the C-terminal catalytic domains. The two PfCLKs form complexes with proteins with predicted nuclease
Nuclease
A nuclease is an enzyme capable of cleaving the phosphodiester bonds between the nucleotide subunits of nucleic acids. Older publications may use terms such as "polynucleotidase" or "nucleodepolymerase"....
, phosphatase
Phosphatase
A phosphatase is an enzyme that removes a phosphate group from its substrate by hydrolysing phosphoric acid monoesters into a phosphate ion and a molecule with a free hydroxyl group . This action is directly opposite to that of phosphorylases and kinases, which attach phosphate groups to their...
or helicase
Helicase
Helicases are a class of enzymes vital to all living organisms. They are motor proteins that move directionally along a nucleic acid phosphodiester backbone, separating two annealed nucleic acid strands using energy derived from ATP hydrolysis.-Function:Many cellular processes Helicases are a...
functions.
Although the kinases are primarily localized in the parasite nucleus, PfCLK-2 is also present in the cytoplasm. They are important for completion of the asexual replication cycle. Substrates phosphorylated by the PfCLKs include the Sky1p substrate, splicing factor Npl3p, and the plasmodial alternative splicing factor PfASF-1.
Within the genome is a family of four protein kinases (Pfnek-1 to -4) that are related to the NIMA (never-in-mitosis) family of kinases. The members of this latter family play important roles in mitosis
Mitosis
Mitosis is the process by which a eukaryotic cell separates the chromosomes in its cell nucleus into two identical sets, in two separate nuclei. It is generally followed immediately by cytokinesis, which divides the nuclei, cytoplasm, organelles and cell membrane into two cells containing roughly...
and meiosis
Meiosis
Meiosis is a special type of cell division necessary for sexual reproduction. The cells produced by meiosis are gametes or spores. The animals' gametes are called sperm and egg cells....
. Pfnek-1 is expressed in asexual parasites and male gametocytes. It is an essential gene for completion of the asexual cycle. The other three (Pfnek-2, -3 and -4) are expressed predominantly in gametocytes.
A mitogen activated protein kinase (MAP kinase) gene is located on is located on chromosome 14. It is predominantly expressed in gametocytes and gametes/zygotes. The protein has 882 amino acid residues and posses a TDY dual phosphorylation site upstream of the highly conserved VATRWYRAPE sequence within subdomain VIII. Within the carboxyl-terminal segment the protein contains an unusually large and highly charged domain. This region includes two repetitive sequences of either a tetrapeptide or octapeptide motif.
A subgroup of cyclin-dependent kinases (CDK) including crk-5 have an activation loop that contains a novel Proline
Proline
Proline is an α-amino acid, one of the twenty DNA-encoded amino acids. Its codons are CCU, CCC, CCA, and CCG. It is not an essential amino acid, which means that the human body can synthesize it. It is unique among the 20 protein-forming amino acids in that the α-amino group is secondary...
-Threonine
Threonine
Threonine is an α-amino acid with the chemical formula HO2CCHCHCH3. Its codons are ACU, ACA, ACC, and ACG. This essential amino acid is classified as polar...
-x-Cytosine
Cytosine
Cytosine is one of the four main bases found in DNA and RNA, along with adenine, guanine, and thymine . It is a pyrimidine derivative, with a heterocyclic aromatic ring and two substituents attached . The nucleoside of cytosine is cytidine...
motif which is absent from all known CDKs outside the Apicomplexa.
The plasmodial mitogen-activated protein kinase kinase Pfnek3 has both serine
Serine
Serine is an amino acid with the formula HO2CCHCH2OH. It is one of the proteinogenic amino acids. By virtue of the hydroxyl group, serine is classified as a polar amino acid.-Occurrence and biosynthesis:...
/threonine
Threonine
Threonine is an α-amino acid with the chemical formula HO2CCHCHCH3. Its codons are ACU, ACA, ACC, and ACG. This essential amino acid is classified as polar...
and tyrosine kinase
Tyrosine kinase
A tyrosine kinase is an enzyme that can transfer a phosphate group from ATP to a protein in a cell. It functions as an "on" or "off" switch in many cellular functions....
activities.
Cysteine proteases
A number of cysteine proteases have been identified this organism including four falcipains, serine repeat antigens (SERA), dipeptidyl aminopeptidase 1, dipeptidyl aminopeptidase 3 and a calpainCalpain
A calpain is a protein belonging to the family of calcium-dependent, non-lysosomal cysteine proteases expressed ubiquitously in mammals and many other organisms. Calpains constitute the C2 family of protease clan CA in the MEROPS database...
homolog. The falcipains belong to the papain
Papain
Papain, also known as papaya proteinase I, is a cysteine protease enzyme present in papaya and mountain papaya .-Papain family:...
family of enzymes. Falcipain-1 appears to be important in the development of the oocysts in the mosquito. Falcipain-2 is involved in the hydrolysis of haemoglobin and appears to be a non essential gene. Falcipain-3 appears to be an essential gene but its function has yet to be firmly established. Dipeptidyl aminopeptidase 1 is found in the food vacuole and is also an essential gene. Dipeptidyl aminopeptidase 3 appears to be involved in the release of the merozoites from the erythrocyte. SERA-5 and SERA-6 appear to be essential genes: SERA-5 also seems to be involved with egress of the merozoites.
Metallopeptidases
There is at least one M1 family aminopeptidase in the genome (PfA-M1). This is a zinc binding metalopeptidase with optimal activity at pH 7.4, and remains at least 40% active between pH 5.8-8.6. Immunofluorescence studies have shown that in trophozoites that it diffusely found in the parasite cytoplasm with accumulations outside the digestive vacuole while in schizonts it is progressively located to a vesicle like pattern ending as a single location in released merozoites. It exists as two major isoforms, a nuclear 120 kDa species and a processed species consisting of a complex of 68 and 35 kDa fragments.There are at least 2 essential metallopeptidases encoded in the genome - PfA-M1 and Pf-LAP. Specific inhibition of PfA-M1 causes swelling of the parasite digestive vacuole and prevented proteolysis of haemoglobin derived oligopeptides. This inhibition is lethal to the parasite probably by starvation. Inhibition of Pf-LAP is lethal to parasites early in the life cycle, prior to the onset of haemoglobin degradation suggesting a different role for this enzyme.
Other proteases
At least four aspartyl proteaseProtease
A protease is any enzyme that conducts proteolysis, that is, begins protein catabolism by hydrolysis of the peptide bonds that link amino acids together in the polypeptide chain forming the protein....
s known as plasmepsins are involved in the degradation of hemoglobin
Hemoglobin
Hemoglobin is the iron-containing oxygen-transport metalloprotein in the red blood cells of all vertebrates, with the exception of the fish family Channichthyidae, as well as the tissues of some invertebrates...
by Plasmodium falciparum.
The histo-aspartic protease (HAP) has been crystalised. This protein has high sequence similarity to pepsin
Pepsin
Pepsin is an enzyme whose precursor form is released by the chief cells in the stomach and that degrades food proteins into peptides. It was discovered in 1836 by Theodor Schwann who also coined its name from the Greek word pepsis, meaning digestion...
-like aspartic
Aspartic acid
Aspartic acid is an α-amino acid with the chemical formula HOOCCHCH2COOH. The carboxylate anion, salt, or ester of aspartic acid is known as aspartate. The L-isomer of aspartate is one of the 20 proteinogenic amino acids, i.e., the building blocks of proteins...
proteases, but one of the two catalytic aspartates, Asp32, is replaced in this enzyme by a histidine
Histidine
Histidine Histidine, an essential amino acid, has a positively charged imidazole functional group. It is one of the 22 proteinogenic amino acids. Its codons are CAU and CAC. Histidine was first isolated by German physician Albrecht Kossel in 1896. Histidine is an essential amino acid in humans...
residue. The propeptide interacts with the C-terminal domain of the enzyme, forcing the N- and C- terminal domains apart. This mechanically separates His32 and Asp215 and prevents formation of the mature active site. This mechanism is similar to those of other proplasmepsins. The enzyme has a number of unique features and may be a useful drug target.
A SUMO
SUMO protein
Small Ubiquitin-like Modifier or SUMO proteins are a family of small proteins that are covalently attached to and detached from other proteins in cells to modify their function...
specific protease PfSENP1 (PFL1635w) has been identified in the genome but its importance if any is not known
Erythrocyte proteins taken up
A small number of erythrocytic proteins are taken up by the parasite during the course of its life cycle. The role these play is not clear. Among these proteins is dematin which interacts with the parasite's 14-3-3 protein.The parasite is capable of making use of the erythrocyte's own enzymes. The enzymes PAK1
PAK1
Serine/threonine-protein kinase PAK 1 is an enzyme that in humans is encoded by the PAK1 gene.-Interactions:PAK1 has been shown to interact with C-Raf, NCK1, LIMK1, RAC1, ARHGEF2, BMX, ARPC1B, DYNLL1, Cyclin-dependent kinase 5, PAK1IP1 and CDC42....
and MEK1 neither of which are encoded in the Plasmodium genome have been shown to be phosphorylated and activated during the course of infection' In vitro work has shown that inhibition of these enzymes is fatal to the parasite.
Transport/secretion
Both the cytoplasmic pH (7.3) and the inside-negative plasma membrane potential (-95mV) are kept fairly constant during the intra erythrocytic cycle. This is due to the action of a V-type H(+)-ATPase which is also responsible for the pH of the digestive vacuole.The intracellular concentration of chloride ions has been estimated to be 48 milliMolar. It appears to actively import using ATP
Adenosine triphosphate
Adenosine-5'-triphosphate is a multifunctional nucleoside triphosphate used in cells as a coenzyme. It is often called the "molecular unit of currency" of intracellular energy transfer. ATP transports chemical energy within cells for metabolism...
both hydrogen
Hydrogen
Hydrogen is the chemical element with atomic number 1. It is represented by the symbol H. With an average atomic weight of , hydrogen is the lightest and most abundant chemical element, constituting roughly 75% of the Universe's chemical elemental mass. Stars in the main sequence are mainly...
ion
Ion
An ion is an atom or molecule in which the total number of electrons is not equal to the total number of protons, giving it a net positive or negative electrical charge. The name was given by physicist Michael Faraday for the substances that allow a current to pass between electrodes in a...
s and chloride
Chloride
The chloride ion is formed when the element chlorine, a halogen, picks up one electron to form an anion Cl−. The salts of hydrochloric acid HCl contain chloride ions and can also be called chlorides. The chloride ion, and its salts such as sodium chloride, are very soluble in water...
ions in a linked fashion via a DIDS sensitive transporter in the cytoplasmic membrane.
One difficulty the parasite has in acquiring nutrients from the cytoplasm is the presence of phosphate groups on these molecules. It appears to have overcome this by secreting an acid phosphatase (glideosome-associated protein 50 - GAP50 ) into the cytoplasm that is then taken up into the food vacuole.
The parasite is unable to synthesize purines (including adenosine, hypoxanthine
Hypoxanthine
Hypoxanthine is a naturally occurring purine derivative. It is occasionally found as a constituent of nucleic acids where it is present in the anticodon of tRNA in the form of its nucleoside inosine. It has a tautomer known as 6-Hydroxypurine. Hypoxanthine is a necessary additive in certain cell,...
and adenine
Adenine
Adenine is a nucleobase with a variety of roles in biochemistry including cellular respiration, in the form of both the energy-rich adenosine triphosphate and the cofactors nicotinamide adenine dinucleotide and flavin adenine dinucleotide , and protein synthesis, as a chemical component of DNA...
) and must take these up from the host. Three purine transporters have been studied: the human equilibrative nucleoside transporter
Equilibrative nucleoside transporter
The equilibrative nucleoside transporter family, also known as SLC29, is a group of plasmalemmal transport proteins which transport nucleoside substrates like adenosine into cells. There are four known ENTs, designated ENT1, ENT2, ENT3, and ENT4...
(hENT1), the human facilitative nucleobase transporter (hFNT1) and the parasite-induced new permeation pathway (NPP). The bulk of transport is facilitated by host's own transporters rather than through the NPP. Hypoxanthine and adenine were transported mainly through the hFNT1 pathway whereas adenosine entered predominantly through the hENT1 system. The rate of purine uptake in infected cells was approximately twice that of uninfected erythrocytes. The rate of adenosine uptake was greater than the rate of hypoxanthine uptake in infected human red blood cells. Furosemide
Furosemide
Furosemide or frusemide is a loop diuretic used in the treatment of congestive heart failure and edema. It is most commonly marketed by Sanofi-Aventis under the brand name Lasix...
inhibits the transport of purine bases through the hFNT1.
The parasite possesses its own Equilibrative Nucleoside Transporter 1. All members of this protein family have 11 transmembrane segments. The gene product is located in the parasite's plasma membrane and knock out mutants have shown that this is an essential gene at least at physiological concentrations. In the 11th transmembrane segment two mutations have been shown to affect its activity: a phenylalanine
Phenylalanine
Phenylalanine is an α-amino acid with the formula C6H5CH2CHCOOH. This essential amino acid is classified as nonpolar because of the hydrophobic nature of the benzyl side chain. L-Phenylalanine is an electrically neutral amino acid, one of the twenty common amino acids used to biochemically form...
(Phe) to leucine
Leucine
Leucine is a branched-chain α-amino acid with the chemical formula HO2CCHCH2CH2. Leucine is classified as a hydrophobic amino acid due to its aliphatic isobutyl side chain. It is encoded by six codons and is a major component of the subunits in ferritin, astacin and other 'buffer' proteins...
(Leu) at residue 394 (F394L) via cytosine
Cytosine
Cytosine is one of the four main bases found in DNA and RNA, along with adenine, guanine, and thymine . It is a pyrimidine derivative, with a heterocyclic aromatic ring and two substituents attached . The nucleoside of cytosine is cytidine...
(C) or uracil
Uracil
Uracil is one of the four nucleobases in the nucleic acid of RNA that are represented by the letters A, G, C and U. The others are adenine, cytosine, and guanine. In RNA, uracil binds to adenine via two hydrogen bonds. In DNA, the uracil nucleobase is replaced by thymine.Uracil is a common and...
(U) to adenosine (A) or guanine
Guanine
Guanine is one of the four main nucleobases found in the nucleic acids DNA and RNA, the others being adenine, cytosine, and thymine . In DNA, guanine is paired with cytosine. With the formula C5H5N5O, guanine is a derivative of purine, consisting of a fused pyrimidine-imidazole ring system with...
(G) at the third codon position and a cysteine
Cysteine
Cysteine is an α-amino acid with the chemical formula HO2CCHCH2SH. It is a non-essential amino acid, which means that it is biosynthesized in humans. Its codons are UGU and UGC. The side chain on cysteine is thiol, which is polar and thus cysteine is usually classified as a hydrophilic amino acid...
(Cys) to glycine
Glycine
Glycine is an organic compound with the formula NH2CH2COOH. Having a hydrogen substituent as its 'side chain', glycine is the smallest of the 20 amino acids commonly found in proteins. Its codons are GGU, GGC, GGA, GGG cf. the genetic code.Glycine is a colourless, sweet-tasting crystalline solid...
(Gly) mutation at either glycine in a conserved glycine-X-X-glycine motif (where X is any amino acid) via a cytosine to uracil at the second codon position. Additional work suggests that the 11th transmembrane segment is largely alpha helical. It has been suggested that this transmembrane segment may be the actual purine transport channel.
The parasite has an absolute requirement for isoleucine
Isoleucine
Isoleucine is an α-amino acid with the chemical formula HO2CCHCHCH2CH3. It is an essential amino acid, which means that humans cannot synthesize it, so it must be ingested. Its codons are AUU, AUC and AUA....
- an amino acid
Amino acid
Amino acids are molecules containing an amine group, a carboxylic acid group and a side-chain that varies between different amino acids. The key elements of an amino acid are carbon, hydrogen, oxygen, and nitrogen...
absent from human haemoglobin. A saturable neutral amino acid (methionine
Methionine
Methionine is an α-amino acid with the chemical formula HO2CCHCH2CH2SCH3. This essential amino acid is classified as nonpolar. This amino-acid is coded by the codon AUG, also known as the initiation codon, since it indicates mRNA's coding region where translation into protein...
, leucine
Leucine
Leucine is a branched-chain α-amino acid with the chemical formula HO2CCHCH2CH2. Leucine is classified as a hydrophobic amino acid due to its aliphatic isobutyl side chain. It is encoded by six codons and is a major component of the subunits in ferritin, astacin and other 'buffer' proteins...
, isoleucine) transporter appears to be encoded by the parasite and this protein functions in the infected erythrocyte membrane.
The clag3 on chromosome 3 appear to be involved in anion transport rather than in cell adherence as originally thought.
The P. falciparum NA+/H+ exchanger (PfNHE1) is located on chromosome 13 (gene PF13_0019).
Two folate transporters (PfFT1 and PfTF2) have been cloned. Substrates include folic acid
Folic acid
Folic acid and folate , as well as pteroyl-L-glutamic acid, pteroyl-L-glutamate, and pteroylmonoglutamic acid are forms of the water-soluble vitamin B9...
, folinic acid
Folinic acid
Folinic acid or leucovorin , generally administered as calcium or sodium folinate , is an adjuvant used in cancer chemotherapy involving the drug methotrexate. It is also used in synergistic combination with the chemotherapy agent 5-fluorouracil.Levofolinic acid and its salts are the enantiopure...
, the folate precursor pABA and the human folate catabolite pABAG(n). 5-methyl tetrahydofolate is not transported by PfFT1 and only poorly by PfFT2. The activity of both transporters may be inhibited by probenecid
Probenecid
Probenecid is a uricosuric drug that increases uric acid excretion in the urine. It is primarily used in treating gout and hyperuricemia.Probenecid was developed as an alternative to caronamide...
or methotrexate
Methotrexate
Methotrexate , abbreviated MTX and formerly known as amethopterin, is an antimetabolite and antifolate drug. It is used in treatment of cancer, autoimmune diseases, ectopic pregnancy, and for the induction of medical abortions. It acts by inhibiting the metabolism of folic acid. Methotrexate...
.
An intracellular purine
Purine
A purine is a heterocyclic aromatic organic compound, consisting of a pyrimidine ring fused to an imidazole ring. Purines, including substituted purines and their tautomers, are the most widely distributed kind of nitrogen-containing heterocycle in nature....
permease
Permease
The permeases are membrane transport proteins, a class of multipass transmembrane proteins that facilitate the diffusion of a specific molecule in or out of the cell by passive transport...
(PfNT2) has been shown to be localised to the endoplasmic reticulum
Endoplasmic reticulum
The endoplasmic reticulum is an organelle of cells in eukaryotic organisms that forms an interconnected network of tubules, vesicles, and cisternae...
. This protein is a member of the equilibrative nucleoside transporter family.
The clag3 gene family encode a parasite ion channel known as the plasmodial surface anion channel. Its activation appears to involve an intracellular domain.
Actin
Within the genome are encoded two forms of the protein actinActin
Actin is a globular, roughly 42-kDa moonlighting protein found in all eukaryotic cells where it may be present at concentrations of over 100 μM. It is also one of the most highly-conserved proteins, differing by no more than 20% in species as diverse as algae and humans...
- I and II. The first form (I) is present in significantly greater quantities. Actin II appears to be essential for the process of exflagellation. Deletion of this gene results in viable asexual stages. During the formation of the male gametes actin I is found initially in both the nucleus and the cytoplasm. After activation it is found only in the cytoplasm. In actin II deletion mutants actin I remains in both the nucleus and the cytoplasm after activation. Morphologically in the actin II mutants male gametocyte DNA was replicates normally and axonemes are assembled but egress from the host cell is inhibited and axoneme motility is abolished.
Two proteins P. falciparum actin-depolymerizing factor 1 (PfADF1) and P. falciparum actin-depolymerizing factor 2 (PfADF2) are involved in the polymerisation of actin. PfADF1 has ben crystalised and despite having significant differences from other proteins with similar function it is capable of severing actin filaments. PfADF2, like canonical ADF proteins but unlike ADF1, binds to both globular and filamentous actin, severing the filaments and inducing nucleotide exchange on the actin monomer. The crystal structure of PfADF1 shows major differences from the ADF consensus, explaining the lack of F-actin binding. PfADF2 structurally resembles the canonical members of the ADF/cofilin family.
The actins found In Plasmodium and in Toxoplasma are divergent both in sequence and function and only form short, unstable filaments in contrast to the stability of conventional actin filaments. This inherent instability of parasite's actin filaments is a critical adaptation for their gliding motility.
Actin is involved in the expression of the var genes. The var introns interact with an 18 base pair nuclear protein binding element which recruits actin and repositions the var DNA from a transcriptionally repressive to a transcriptionally active perinuclear compartment.
Surface exposed proteins
EnolaseEnolase
Enolase, also known as phosphopyruvate dehydratase, is a metalloenzyme responsible for the catalysis of the conversion of 2-phosphoglycerate to phosphoenolpyruvate , the ninth and penultimate step of glycolysis. Enolase belongs to the class Lyase. Enolase can also catalyze the reverse reaction,...
is bound to the surface of P. falciparum and several other pathogens. In this location it binds plasminogen which is thought to function in the degradation of the extracellular matrix surrounding the targeted host cell, thereby facilitating pathogen invasion.
The ETRAMP family is characterized by a predicted signal peptide
Signal peptide
A signal peptide is a short peptide chain that directs the transport of a protein.Signal peptides may also be called targeting signals, signal sequences, transit peptides, or localization signals....
, a short lysine
Lysine
Lysine is an α-amino acid with the chemical formula HO2CCH4NH2. It is an essential amino acid, which means that the human body cannot synthesize it. Its codons are AAA and AAG....
rich stretch, an internal transmembrane domain and a highly charged C-terminal region of variable length. The highly charged terminal region appears to be involved in protein-protein interactions. The gene ETRAMP 10.3 has been shown to be expressed in the liver, sporozoites and blood stages. Within the liver and blood stages it is localized to the parasitophorous vacuole membrane. It is also exported to the erythrocyte during the blood stages. It appears to be an essential gene in the blood stages.
The receptor for the attachment protein PfRh4 has been identified as complement receptor 1
Complement receptor 1
Erythrocyte complement receptor 1 is a human gene....
.
The TRAP like protein is invovled in cell motility.
The circumsporozoite- and thrombospondin-related adhesive protein (CTRP) is a modular multidomain protein containing six tandem von Willebrand factor
Von Willebrand factor
von Willebrand factor is a blood glycoprotein involved in hemostasis. It is deficient or defective in von Willebrand disease and is involved in a large number of other diseases, including thrombotic thrombocytopenic purpura, Heyde's syndrome, and possibly hemolytic-uremic syndrome...
A like domains and seven tandem thrombospondin
Thrombospondin
Thrombospondins are secreted proteins with antiangiogenic abilities. TSP was discovered by Nancy L. Baenziger.-Types:The thrombospondins are a family of multifunctional proteins...
type I repeat-like domains. The A domains of CTRP are critical for ookinete gliding motility and oocyst formation. The thrombospondin domains are fully redundant.
The cell-traversal protein for ookinetes and sporozoites (CelTOS) is a protein involved in the invasion of both vertebrate and insect host cells.
Merozoite surface protein 7 appears to enhance the virulence of the parasite at least in the rodent.
Merozoite surface protein
Merozoite surface protein
A merozoite surface protein is a protein molecule taken from the skin, or surface, of a merozoite. A merozoite is a "daughter cell" of a protozoan.Merozoite surface proteins, or MSPs, are useful in researching malaria, a disease caused by protozoans....
2 is one of the most abundant proteins on the surface of merozoites, is intrinsically unstructured and forms amyloid
Amyloid
Amyloids are insoluble fibrous protein aggregates sharing specific structural traits. Abnormal accumulation of amyloid in organs may lead to amyloidosis, and may play a role in various neurodegenerative diseases.-Definition:...
-like fibrils in solution.
The mature parasite-infected erythrocyte surface antigen (MESA) is exported to the erythrocyte cytoplasm where it binds to the N-terminal 30 kiloDalton domain of the erythrocyte protein 4.1R via a 19-residue sequence. This sequence is also found in a number of other proteins in the parasite. Their role in remodeling of the erythrocyte are still under investigation.
The proteins Pf12, Pf34, Pf92 and Pf38 are associated with detergent resistant membrane microdomains through glycosylphosphatidylinositol anchor sequences. These microdomains are considered organizing centers for the assembly of molecules implicated in cell signaling.
Positive diversifying selection is present in clag2, clag8 and clag9 but not in clag3.1 and clag3.2.
Ubiquitin
The addition of the small protein ubiquitinUbiquitin
Ubiquitin is a small regulatory protein that has been found in almost all tissues of eukaryotic organisms. Among other functions, it directs protein recycling.Ubiquitin can be attached to proteins and label them for destruction...
to other proteins as part of post translational processing is widespread in most eukaryotes. This is also the case with P. falciparum where this process occurs at all stages of the asexual life cycle.
Gene regulation
CCR4-associated factor 1 is involved in the regulation more than 1000 genes during malaria parasite's intraerythrocytic stages. Mutations in this gene result in mistimed expression, aberrant accumulation and localization of proteins involved in parasite egress and invasion of new host cells. This leads to the premature release of predominantly half-finished merozoites in turn drastically reducing the intraerythrocytic growth rate of the parasite.A homolog of the DEAD box
DEAD box
DEAD box proteins are involved in an assortment of metabolic processes that involve RNA. They are highly conserved in nine domains and can be found in both prokaryotes and eukaryotes, but not all...
(Asparagine
Asparagine
Asparagine is one of the 20 most common natural amino acids on Earth. It has carboxamide as the side-chain's functional group. It is not an essential amino acid...
-Glutamate-Alanine
Alanine
Alanine is an α-amino acid with the chemical formula CH3CHCOOH. The L-isomer is one of the 20 amino acids encoded by the genetic code. Its codons are GCU, GCC, GCA, and GCG. It is classified as a nonpolar amino acid...
-Asparagine) RNA
RNA
Ribonucleic acid , or RNA, is one of the three major macromolecules that are essential for all known forms of life....
helicase
Helicase
Helicases are a class of enzymes vital to all living organisms. They are motor proteins that move directionally along a nucleic acid phosphodiester backbone, separating two annealed nucleic acid strands using energy derived from ATP hydrolysis.-Function:Many cellular processes Helicases are a...
DDX19 (Dbp5) has been cloned from the P. falciparum genome and has been termed PfD66. This protein has intrinsic nucleic acid dependent ATPase
ATPase
ATPases are a class of enzymes that catalyze the decomposition of adenosine triphosphate into adenosine diphosphate and a free phosphate ion. This dephosphorylation reaction releases energy, which the enzyme harnesses to drive other chemical reactions that would not otherwise occur...
and RNA binding activities and ATP dependent bipolar DNA
DNA
Deoxyribonucleic acid is a nucleic acid that contains the genetic instructions used in the development and functioning of all known living organisms . The DNA segments that carry this genetic information are called genes, but other DNA sequences have structural purposes, or are involved in...
and RNA unwinding activity.
SAP1 has been shown to be involved in the post transcriptional control of liver
Liver
The liver is a vital organ present in vertebrates and some other animals. It has a wide range of functions, including detoxification, protein synthesis, and production of biochemicals necessary for digestion...
stage genes.
The protein PfMyb1 is a transcription factor belonging to the tryptophan
Tryptophan
Tryptophan is one of the 20 standard amino acids, as well as an essential amino acid in the human diet. It is encoded in the standard genetic code as the codon UGG...
cluster family. Inhibition of this gene reduces growth by ~40% with the mortality being concentrated at the trophozoite-schizont interface.
Heat shock proteins
A number of heat shock proteinHeat shock protein
Heat shock proteins are a class of functionally related proteins involved in the folding and unfolding of other proteins. Their expression is increased when cells are exposed to elevated temperatures or other stress. This increase in expression is transcriptionally regulated...
s 40 (hsp40) have been predicted from the sequenced genome. Only one is predicted to be a cytosolic canonical Hsp40 capable of interacting with the major cytosolic Hsp70 an interaction that has been confirmed experimentally.
PfGECO is a type IV heat shock protein 40 expressed in gametocyte stages I to IV and is exported to the erythrocyte cytoplasm. This gene appears to be non essential.
Heat shock proteins Hsp70 and Hsp90 are both expressed in P. falciparum. They are linked by an essential adaptor protein known as the Hsp70-Hsp90 organising protein (Hop). This protein co-localises with PfHsp70 and PfHsp90 at the trophozoite stage and forms a complex with them.
The protein Aha1 interacts with HSP90.
Miscellaneous proteins
Polyamine biosynthesis in these parasites is controlled by a unique bifunctional S-adenosylmethionine decarboxylase/ornithine decarboxylaseOrnithine decarboxylase
The enzyme ornithine decarboxylase catalyzes the decarboxylation of ornithine to form putrescine. This reaction is the committed step in polyamine synthesis. In humans, this protein has 461 amino acids and forms a homodimer....
(PfAdoMetDC/ODC). On the secondary structure level PfAdoMetDC is similar to that of the human protein. This bifunctional enzyme ensure coordination decarboxylated AdoMet and putrescine
Putrescine
Putrescine is a foul-smelling organic chemical compound NH24NH2 that is related to cadaverine; both are produced by the breakdown of amino acids in living and dead organisms and both are toxic in large doses...
for the subsequent synthesis of spermidine
Spermidine
Spermidine is a polyamine involved in cellular metabolism. Its known actions include:#Inhibition of neuronal nitric oxide synthase, nNOS#Assisting the in vitro process of transcribing RNA via stimulation of T4 polynucleotide kinase and T7 RNA polymerase activity; it binds to and precipitates DNA...
.
P. falciparum contains both cytosolic and mitochondrial serine
Serine
Serine is an amino acid with the formula HO2CCHCH2OH. It is one of the proteinogenic amino acids. By virtue of the hydroxyl group, serine is classified as a polar amino acid.-Occurrence and biosynthesis:...
hydroxymethyltransferase isoforms.
The first two reactions of the pentose phosphate pathway
Pentose phosphate pathway
The pentose phosphate pathway is a process that generates NADPH and pentoses . There are two distinct phases in the pathway. The first is the oxidative phase, in which NADPH is generated, and the second is the non-oxidative synthesis of 5-carbon sugars...
in P. falciparum are catalysed by a single bifunctional enzyme - glucose 6-phosphate dehydrogenase 6-phosphogluconolactonase
6-phosphogluconolactonase
6-Phosphogluconolactonase is an enzyme in the pentose phosphate pathway. It converts 6-phosphogluconolactone to 6-phosphogluconate....
. This is distinct from the case in humans where the enzymes are separate. In animal
Animal
Animals are a major group of multicellular, eukaryotic organisms of the kingdom Animalia or Metazoa. Their body plan eventually becomes fixed as they develop, although some undergo a process of metamorphosis later on in their life. Most animals are motile, meaning they can move spontaneously and...
s this pathway is usually found in the cytosol while in plant
Plant
Plants are living organisms belonging to the kingdom Plantae. Precise definitions of the kingdom vary, but as the term is used here, plants include familiar organisms such as trees, flowers, herbs, bushes, grasses, vines, ferns, mosses, and green algae. The group is also called green plants or...
s it is found in the plastid
Plastid
Plastids are major organelles found in the cells of plants and algae. Plastids are the site of manufacture and storage of important chemical compounds used by the cell...
s. The location of this reaction is not currently known in P. falciparum.
Fusions between these two enzymes (glucose 6-phosphate dehydrogenase and 6-phosphogluconolactonase
6-phosphogluconolactonase
6-Phosphogluconolactonase is an enzyme in the pentose phosphate pathway. It converts 6-phosphogluconolactone to 6-phosphogluconate....
) have also been reported in chordates. The chordate fusion differs in its orientation to that in Plasmodium (in Plasmodium the 6-phosphogluconolactone is found at the N-terminus of the glucose 6-phosphate dehydrogenase protein) indicating that at least two separate fusion events have occurred. The metazoan fusion appears to have occurred near the bases of the metazoan and apicomplexa
Apicomplexa
The Apicomplexa are a large group of protists, most of which possess a unique organelle called apicoplast and an apical complex structure involved in penetrating a host's cell. They are unicellular, spore-forming, and exclusively parasites of animals. Motile structures such as flagella or...
n lineages. This fusion event was not found in any of the three sequenced Cryptosporidium
Cryptosporidium
Cryptosporidium is a protozoan that can cause gastro-intestinal illness with diarrhea in humans.Cryptosporidium is the organism most commonly isolated in HIV positive patients presenting with diarrhea...
genomes. It was not found in Perkinsus marinus
Perkinsus marinus
Perkinsus marinus is a prevalent pathogen of oysters, causing massive mortality in oyster populations. The disease it causes is known as "Dermo" ,and is characterized by proteolytic degradation of oyster tissues...
or in either of the ciliate (Paramecium tetraurelia and Tetrahymena thermophila) genomes. More data will be needed to estimate the timing of this fusion event.
Only one of the one of the two metazoan paralogs of glucose 6-phosphate dehydrogenase is fused indicating that the fusion occurred after a duplication event. This duplication event occurred in an ancestor of the choanoflagellate
Choanoflagellate
The choanoflagellates are a group of free-living unicellular and colonial flagellate eukaryotes considered to be the closest living relatives of the animals...
s and metazoans. Another fusion event between these enzymes occurred in an ancestor of the protozoan parasites Trichomonas
Trichomonas
Trichomonas is a genus of anaerobic protists that are parasites of vertebrates. They are included with the parabasalids.Species of Trichomonas include:*Trichomonas vaginalis, an organism generally living inside the vagina of humans...
and Giardia lamblia
Giardia lamblia
Giardia lamblia is a flagellated protozoan parasite that colonizes and reproduces in the small intestine, causing giardiasis. The giardia parasite attaches to the epithelium by a ventral adhesive disc, and reproduces via binary fission...
. In Giardia the proteins are fused in opposite orientations. A third fusion event occurred between glucose 6-phosphate dehydrogenase with phosphogluconate dehydrogenase
Phosphogluconate dehydrogenase
Phosphogluconate dehydrogenase is an enzyme in the pentose phosphate pathway. It forms ribulose 5-phosphate from 6-phosphogluconate.It is an oxidative carboxylase that catalyses the decarboxylating reduction of 6-phosphogluconate into ribulose 5-phosphate in the presence of NADP. This reaction is a...
in a diatom
Diatom
Diatoms are a major group of algae, and are one of the most common types of phytoplankton. Most diatoms are unicellular, although they can exist as colonies in the shape of filaments or ribbons , fans , zigzags , or stellate colonies . Diatoms are producers within the food chain...
species (Phaeodactylum tricornutum
Phaeodactylum tricornutum
Phaeodactylum tricornutum is a diatom. It is the only species in the genus Phaeodactylum. Unlike other diatoms P. tricornutum can exist in different morphotypes , and changes in cell shape can be stimulated by environmental conditions. This feature can be used to explore the molecular basis of cell...
).
Phosphoinositide-specific phospholipase
Phospholipase
A phospholipase is an enzyme that hydrolyzes phospholipids into fatty acids and other lipophilic substances. There are four major classes, termed A, B, C and D, distinguished by the type of reaction which they catalyze:*Phospholipase A...
C (PI-PLC) is a major regulator of calcium
Calcium
Calcium is the chemical element with the symbol Ca and atomic number 20. It has an atomic mass of 40.078 amu. Calcium is a soft gray alkaline earth metal, and is the fifth-most-abundant element by mass in the Earth's crust...
-dependent signal transduction usually by liberation of calcium from intracellular stores through the action of its product, inositol-(1,4,5)-trisphosphate. These genes are found in P. falciparum and appear to be essential. The genes are twice as long as their mammalian counterparts and belong to the delta class of phospholipase C proteins.
The mechanism of action of the triose phosphate isomerase enzyme has been investigated in some detail. The conserved glutamic acid
Glutamic acid
Glutamic acid is one of the 20 proteinogenic amino acids, and its codons are GAA and GAG. It is a non-essential amino acid. The carboxylate anions and salts of glutamic acid are known as glutamates...
residue at position 97 is involved in the catalytic proton transfer. Modification of this residue may reduce the rate of catalysis by 9000 fold.
There are two translation elongation factor g
EF-G
EF-G or elongation factor G is one of the prokaryotic elongation factors.-Function:The factor EF-G catalyzes the translocation of the tRNA and mRNA down the ribosome at the end of each round of polypeptide elongation. Homologous to EF-Tu + tRNA, EF-G also binds to the ribosome in its GTP-bound...
proteins encoded in the genome. One is located in the mitochondrion and the second in the plastid. Both appear to be inhibitable with fusidic acid
Fusidic acid
Fusidic acid is a bacteriostatic antibiotic that is often used topically in creams and eyedrops, but may also be given systemically as tablets or injections...
The type II NADH:ubiquinone oxidoreductase
Oxidoreductase
In biochemistry, an oxidoreductase is an enzyme that catalyzes the transfer of electrons from one molecule to another...
has been shown to be redundant in the blood forms but to be essential in the mosquito midgut.
Chorismate synthase
Chorismate synthase
In enzymology, a chorismate synthase is an enzyme that catalyzes the chemical reactionHence, this enzyme has one substrate, 5-O--3-phosphoshikimate, and two products, chorismate and phosphate....
(CS) catalyses the seventh and final step of the shikimate pathway. P. falciparum chorismate synthase (PfCS) is unique in terms of enzymatic behavior, cellular localization and in having two additional amino acid inserts compared to any other CS.
Polynucleotide kinase/phosphatase (PNKP) is a bifunctional enzyme that can phosphorylate the 5'-OH termini and dephosphorylate the 3'-phosphate termini of DNA
DNA
Deoxyribonucleic acid is a nucleic acid that contains the genetic instructions used in the development and functioning of all known living organisms . The DNA segments that carry this genetic information are called genes, but other DNA sequences have structural purposes, or are involved in...
. It is a DNA repair enzyme involved in the processing of strand break termini, which permits subsequent repair proteins to replace missing nucleotides and rejoin broken strands. A P. falciparum gene encoding a protein with 24% homology to human PNKP has been cloned. This enzyme dephosphorylates single-stranded substrates or double-stranded substrates with a short 3'-single-stranded overhang, but not double-stranded substrates that mimicked single-strand breaks.
Sir2A is a member of the sirtuin
Sirtuin
Sirtuin or Sir2 proteins are a class of proteins that possess either histone deacetylase or mono-ribosyltransferase activity. Sirtuins regulate important biological pathways in bacteria, archaea and eukaryotes...
family of nicotinamide adenine dinucleotide
Nicotinamide adenine dinucleotide
Nicotinamide adenine dinucleotide, abbreviated NAD, is a coenzyme found in all living cells. The compound is a dinucleotide, since it consists of two nucleotides joined through their phosphate groups. One nucleotide contains an adenine base and the other nicotinamide.In metabolism, NAD is involved...
dependent deacetylases. In P. falciparum it has been has been shown to regulate the expression of surface antigens to evade the detection by host immune surveillance. While it is a poor deacetylator of histone
Histone
In biology, histones are highly alkaline proteins found in eukaryotic cell nuclei that package and order the DNA into structural units called nucleosomes. They are the chief protein components of chromatin, acting as spools around which DNA winds, and play a role in gene regulation...
s it also catalyzes the hydrolysis of medium and long chain fatty acyl groups from lysine
Lysine
Lysine is an α-amino acid with the chemical formula HO2CCH4NH2. It is an essential amino acid, which means that the human body cannot synthesize it. Its codons are AAA and AAG....
residues. Proteins are present in P. falciparum with these modifications and these can be removed by can be removed by PfSir2A in vitro. This suggests that this may be its role rather than the deacetylation of histones.
The telomerase
Telomerase
Telomerase is an enzyme that adds DNA sequence repeats to the 3' end of DNA strands in the telomere regions, which are found at the ends of eukaryotic chromosomes. This region of repeated nucleotide called telomeres contains non-coding DNA material and prevents constant loss of important DNA from...
(tert) is a large protein (2518 codons) and has a predicted molecular weight of ~280 kiloDaltons. It has the usual telomerase specific motifs within the N-terminal half of the protein (GQ/N, CP, QFP and T) and reverse transcriptase (RT) specific motifs in the C-terminal half. The N-terminal half is required for efficient binding of the RNA template, defining the 5′ RNA template boundary, multimerization and interactions with associated proteins. The RT domain is essential for the catalytic activity. The protein contains several nuclear localization signals and is found in the nucleolus
Nucleolus
The nucleolus is a non-membrane bound structure composed of proteins and nucleic acids found within the nucleus. Ribosomal RNA is transcribed and assembled within the nucleolus...
.
A histone deacetylase
Histone deacetylase
Histone deacetylases are a class of enzymes that remove acetyl groups from an ε-N-acetyl lysine amino acid on a histone. This is important because DNA is wrapped around histones, and DNA expression is regulated by acetylation and de-acetylation. Its action is opposite to that of histone...
(HDAC1) has been cloned. The protein has 449 amino acid residues and localises to the nucleus. Its molecular weight is 50 kiloDaltons and it is predominantly expressed in mature asexual blood stages and in gametocytes.
A phosphatidylserine decarboxylase
Phosphatidylserine decarboxylase
In enzymology, a phosphatidylserine decarboxylase is an enzyme that catalyzes the chemical reactionHence, this enzyme has one substrate, phosphatidyl-L-serine, and two products, phosphatidylethanolamine and CO2....
has been cloned from the parasite Plasmodium knowlesi
Plasmodium knowlesi
Plasmodium knowlesi is a primate malaria parasite commonly found in Southeast Asia. It causes malaria in long-tailed macaques , but it may also infect humans, either naturally or artificially....
. It seems highly probably this enzyme is also found in P. falciparum.
Tyrosyl tRNA synthetase is secreted by the parasite into the cytoplasm
Cytoplasm
The cytoplasm is a small gel-like substance residing between the cell membrane holding all the cell's internal sub-structures , except for the nucleus. All the contents of the cells of prokaryote organisms are contained within the cytoplasm...
of the infected erythrocyte. On lysis of the erythrocyte it is released into the blood stream where it is pro inflamatory. It is specifically bound by and taken up by host macrophages and leads to enhanced secretion of the cytokine
Cytokine
Cytokines are small cell-signaling protein molecules that are secreted by the glial cells of the nervous system and by numerous cells of the immune system and are a category of signaling molecules used extensively in intercellular communication...
s tumor necrosis factor-alpha and interleukin 6
Interleukin 6
Interleukin-6 is a protein that in humans is encoded by the IL6 gene.IL-6 is an interleukin that acts as both a pro-inflammatory and anti-inflammatory cytokine. It is secreted by T cells and macrophages to stimulate immune response, e.g. during infection and after trauma, especially burns or other...
. This interaction also increases the adherence linked host endothelial receptors ICAM-1
ICAM-1
ICAM-1 also known as CD54 is a protein that in humans is encoded by the ICAM1 gene. This gene encodes a cell surface glycoprotein which is typically expressed on endothelial cells and cells of the immune system...
and VCAM-1
VCAM-1
Vascular cell adhesion protein 1 also known as vascular cell adhesion molecule 1 or cluster of differentiation 106 is a protein that in humans is encoded by the VCAM1 gene...
.
The MutL homolog (MLH) - part of the DNA mismatch repair
DNA mismatch repair
DNA mismatch repair is a system for recognizing and repairing erroneous insertion, deletion and mis-incorporation of bases that can arise during DNA replication and recombination, as well as repairing some forms of DNA damage....
system - has been cloned. MLH possess ATPase
ATPase
ATPases are a class of enzymes that catalyze the decomposition of adenosine triphosphate into adenosine diphosphate and a free phosphate ion. This dephosphorylation reaction releases energy, which the enzyme harnesses to drive other chemical reactions that would not otherwise occur...
and endonuclease
Endonuclease
Endonucleases are enzymes that cleave the phosphodiester bond within a polynucleotide chain, in contrast to exonucleases, which cleave phosphodiester bonds at the end of a polynucleotide chain. Typically, a restriction site will be a palindromic sequence four to six nucleotides long. Most...
activities. Its' expression is maximal in the schizont stage.
Both the 2-C-methyl-d-erythritol-4-phosphate and shikimate pathways are functional in P. falciparum and vitamin E
Vitamin E
Vitamin E is used to refer to a group of fat-soluble compounds that include both tocopherols and tocotrienols. There are many different forms of vitamin E, of which γ-tocopherol is the most common in the North American diet. γ-Tocopherol can be found in corn oil, soybean oil, margarine and dressings...
biosynthesis occurs.
Membrane biogenesis in this organism involves the enzyme phosphoethanolamine methyltransferase which catalyses the methylation of phosphoethanolamine to phosphocholine
Phosphocholine
Phosphocholine is an intermediate in the synthesis of phosphatidylcholine in tissues. Phosphocholine is made in a reaction, catalyzed by choline kinase, that converts ATP + Choline into Phosphocholine and ADP...
. This pathway is found in plant
Plant
Plants are living organisms belonging to the kingdom Plantae. Precise definitions of the kingdom vary, but as the term is used here, plants include familiar organisms such as trees, flowers, herbs, bushes, grasses, vines, ferns, mosses, and green algae. The group is also called green plants or...
s and nematode
Nematode
The nematodes or roundworms are the most diverse phylum of pseudocoelomates, and one of the most diverse of all animals. Nematode species are very difficult to distinguish; over 28,000 have been described, of which over 16,000 are parasitic. It has been estimated that the total number of nematode...
s but not in humans. The enzymes in P. falciparum is a multi-functional unlike that of plants and nematodes. The enzyme from P. falciparum has been cloned and its structure solved.
A homolog of the inhibitor of protein phosphatase 1
Protein phosphatase 1
Phosphoprotein phosphatase 1 belongs to a certain class of phosphatases known as protein serine/ threonine phosphatases. This type of phosphatase includes metal-dependent protein phosphatases and aspartate-based phosphatases...
has been cloned. This gene is essential for survival and appears to be localised to the nucleus. A conserved 41- Lysine
Lysine
Lysine is an α-amino acid with the chemical formula HO2CCH4NH2. It is an essential amino acid, which means that the human body cannot synthesize it. Its codons are AAA and AAG....
-Valine
Valine
Valine is an α-amino acid with the chemical formula HO2CCHCH2. L-Valine is one of 20 proteinogenic amino acids. Its codons are GUU, GUC, GUA, and GUG. This essential amino acid is classified as nonpolar...
-Valine-Arginine
Arginine
Arginine is an α-amino acid. The L-form is one of the 20 most common natural amino acids. At the level of molecular genetics, in the structure of the messenger ribonucleic acid mRNA, CGU, CGC, CGA, CGG, AGA, and AGG, are the triplets of nucleotide bases or codons that codify for arginine during...
-Tryptophan
Tryptophan
Tryptophan is one of the 20 standard amino acids, as well as an essential amino acid in the human diet. It is encoded in the standard genetic code as the codon UGG...
- 45 motif is essential for its inhibition activity.
Genome
The genome of Plasmodium falciparumPlasmodium falciparum
Plasmodium falciparum is a protozoan parasite, one of the species of Plasmodium that cause malaria in humans. It is transmitted by the female Anopheles mosquito. Malaria caused by this species is the most dangerous form of malaria, with the highest rates of complications and mortality...
(clone 3D7) was fully sequenced in 2002. The parasite has a 23 megabase genome, divided into 14 chromosomes. The genome codes for approximately 5,300 genes. About 60% of the putative proteins have little or no similarity to proteins in other organisms, and thus currently have no functional assignment. It is estimated 52.6% of the genome is a coding region, with 53.9% of the putative genes containing at least one intron.
Haploid/diploid
It is haploid during nearly all stages of its life-cycle, except for a brief period after fertilization when it is diploid from the ookinete to sporogenic stages within the mosquitoMosquito
Mosquitoes are members of a family of nematocerid flies: the Culicidae . The word Mosquito is from the Spanish and Portuguese for little fly...
gut.
AT richness
The P. falciparumPlasmodium falciparum
Plasmodium falciparum is a protozoan parasite, one of the species of Plasmodium that cause malaria in humans. It is transmitted by the female Anopheles mosquito. Malaria caused by this species is the most dangerous form of malaria, with the highest rates of complications and mortality...
genome has an AT content of roughly 80.6%. Within the intron and intergenic regions, this AT composition rises to roughly 90%. The putative exons contain an AT content of 76.3%. The parasite's AT content is very high in comparison to other organisms. For example, the entire genomes of Saccharomyces cerevisiae
Saccharomyces cerevisiae
Saccharomyces cerevisiae is a species of yeast. It is perhaps the most useful yeast, having been instrumental to baking and brewing since ancient times. It is believed that it was originally isolated from the skin of grapes...
and Arabidopsis thaliana
Arabidopsis thaliana
Arabidopsis thaliana is a small flowering plant native to Europe, Asia, and northwestern Africa. A spring annual with a relatively short life cycle, arabidopsis is popular as a model organism in plant biology and genetics...
have AT contents of 62% and 65%, respectively.
Translation initiation
This has been examined experimentally for the heat shock protein 86. Like other eukaryotes purines at the -3 and +4 positions are essential for efficient translation. UracilUracil
Uracil is one of the four nucleobases in the nucleic acid of RNA that are represented by the letters A, G, C and U. The others are adenine, cytosine, and guanine. In RNA, uracil binds to adenine via two hydrogen bonds. In DNA, the uracil nucleobase is replaced by thymine.Uracil is a common and...
at the -1 position resulted in 2.5-fold higher reporter activity compared to wild type.
Recombination
The overall recombination rate is 9.6 kilobase per centimorganCentimorgan
In genetics, a centimorgan or map unit is a unit of recombinant frequency for measuring genetic linkage, defined as that distance between chromosome positions for which the expected average number of intervening chromosomal crossovers in a single generation is 0.01. It is often used to infer...
and 54 candidate recombination hotspots have been identified. The centromere
Centromere
A centromere is a region of DNA typically found near the middle of a chromosome where two identical sister chromatids come closest in contact. It is involved in cell division as the point of mitotic spindle attachment...
s are found in chromosome regions largely devoid of recombination activity like other organisms. Within the hotspots a number of motifs were enriched including a 12 base pair G/C-rich motif with 3 base pair periodicity that may interact with a protein containing 11 predicted zinc finger
Zinc finger
Zinc fingers are small protein structural motifs that can coordinate one or more zinc ions to help stabilize their folds. They can be classified into several different structural families and typically function as interaction modules that bind DNA, RNA, proteins, or small molecules...
arrays.
Subtelomeric regions
Throughout the eukaryotic kingdom, the overall structure of chromosome ends is conserved and is characterized by the telomeric tract - a series of short G-rich repeats. This is succeeded by an extensive subtelomeric region consisting of various types and lengths of repeats - the telomere associated sequences (TAS). In general transcription of genes located next to telomeres is repressed, a phenomenon termed the telomere position effect. This effect is somewhat misnamed as it appears to be due to the sequences found in this region rather than the position of the gene.Subtelomeric regions in general are low in gene density, low in transcription, low in recombination, late replicating, are involved in protecting the end from degradation and end-to-end fusions and in completing replication. The subtelomeric repeats can rescue chromosome ends when telomerase
Telomerase
Telomerase is an enzyme that adds DNA sequence repeats to the 3' end of DNA strands in the telomere regions, which are found at the ends of eukaryotic chromosomes. This region of repeated nucleotide called telomeres contains non-coding DNA material and prevents constant loss of important DNA from...
fails, buffer subtelomerically located genes against transcriptional silencing and protect the genome from deleterious rearrangements due to ectopic recombination. They may also be involved in fillers for increasing chromosome size to some minimum threshold level necessary for chromosome stability; act as barriers against transcriptional silencing; provide a location for the adaptive amplification of genes; and be involved in secondary mechanism of telomere maintenance via recombination when telomerase activity is absent. The repressive histone
Histone
In biology, histones are highly alkaline proteins found in eukaryotic cell nuclei that package and order the DNA into structural units called nucleosomes. They are the chief protein components of chromatin, acting as spools around which DNA winds, and play a role in gene regulation...
3 lysine 9 tri-methylation mark and heterochromatin protein 1
Heterochromatin protein 1
The family of Heterochromatin Protein 1 are highly conserved adapter proteins, which have important functions in the cell nucleus...
are found throughout the TAS region and adjacent gene families on all chromosomes. These heterochromatic marks are important in telomere proximal gene silencing.
In parasitic species genes involved in antigenic variation are commonly located in these regions.
The chromsomes of P. falciparum conform to this basic pattern with the ends of the chromosomes consist of a stretch of telomeric GGGTT(T/C)A repeats with an average size of 1.2 kilobases (kb). This is followed by an extensive 20 to 40 kb TAS domain. These show a high degree of conservation within the genome and contain significant amounts of repeated structure. Telomere repeats are followed by a mosaic of six distinct telomere associated repetitive elements (TAREs 1-6) which are always found in the same order but vary in size.
Many genes involved in antigenic variation are located in the subtelomeric regions of the chromosomes. These are divided into the var, rif, and stevor families. Within the genome, there are 59 var, 149 rif, and 28 stevor genes along with multiple pseudogenes and truncations.
The chromosome ends form clusters of 4–7 telomeres that localize around the nuclear periphery. Within this location the telomeric areas undergo frequent recombination which seems to increases antigenic variation.
Transcriptome
A transcriptome analysis has been conducted on the intraerythrocytic development cycle of P. falciparum
Plasmodium falciparum
Plasmodium falciparum is a protozoan parasite, one of the species of Plasmodium that cause malaria in humans. It is transmitted by the female Anopheles mosquito. Malaria caused by this species is the most dangerous form of malaria, with the highest rates of complications and mortality...
. Roughly 60% of the genome is transcriptionally active during this portion of the parasite's life cycle. Whereas many genes appear to have stable mRNA levels throughout the cycle, many of the genes are transcriptionally regulated in a continuous cascade.
The transition from early trophozoite to trophozoite to schizont correlates with the ordered induction of genes related to transcription/translation machinery, metabolic synthesis, energy metabolism, DNA replication, protein degradation, plastid functions, merozoite invasion, and motility.
Closely adjacent genes along the chromosome do not exhibit common transcription characteristics. Thus, genes are likely individually regulated along the parasite chromosome.
Conversely, the apicoplast genome is polycistronic and most of its genes are coexpressed during the intraerythrocytic development cycle.
Introns
The intron splicing has been examined experimentally. The 5' and 3' splice sites agree with the canonical sequences (GT and AG respectively). The 5' consensus motif is weakly conserved and tolerates nucleotide substitution including the fifth nucleotide in the intron. This fifth position, typically a G nucleotide in most eukaryotes, is frequently an A in P. falciparum. The 3' splice site has a strong eukaryotic consensus sequence and a conserved adjacent polypyrimidine tract. The branch point is less well conserved with multiple branch points per intron with some at U instead of the typical A residue. A weak branch point consensus has been identified.Proteome
There are 5,268 predicted proteins in Plasmodium falciparumPlasmodium falciparum
Plasmodium falciparum is a protozoan parasite, one of the species of Plasmodium that cause malaria in humans. It is transmitted by the female Anopheles mosquito. Malaria caused by this species is the most dangerous form of malaria, with the highest rates of complications and mortality...
, and roughly 60% share little or no similarity to proteins in other organisms and thus are without functional assignment. Of the predicted proteins, 31% contain at least one transmembrane domain, and 17.3% have a signal peptide or signal anchor.
It is estimated that 10.4% of the proteome
Proteome
The proteome is the entire set of proteins expressed by a genome, cell, tissue or organism. More specifically, it is the set of expressed proteins in a given type of cells or an organism at a given time under defined conditions. The term is a portmanteau of proteins and genome.The term has been...
is targeted to the apicoplast
Apicoplast
An apicoplast is a derived non-photosynthetic plastid found in most Apicomplexa, including malaria parasites such as Plasmodium falciparum, but not in others such as Cryptosporidium. It originated from an algae through secondary endosymbiosis...
.
It is estimated that 4.7% of the proteome
Proteome
The proteome is the entire set of proteins expressed by a genome, cell, tissue or organism. More specifically, it is the set of expressed proteins in a given type of cells or an organism at a given time under defined conditions. The term is a portmanteau of proteins and genome.The term has been...
is targeted to the mitochondria.
The parasite has different subsets of its proteome expressed during various stages of its developmental cycle. In one study, of the 2,415 proteins were identified in four stages(sporozoite, merozoite, trophozoite, gametocyte), representing 46% of the theoretical number of proteins. Only 6% of the proteins were found in all of the four stages. Of the proteins found, 51% were annotated as hypothetical proteins.
Merozoites contained high levels of cell recognition and invasion proteins. Trophozoites contained proteins implicated in erythrocyte remodeling and hemoglobin
Hemoglobin
Hemoglobin is the iron-containing oxygen-transport metalloprotein in the red blood cells of all vertebrates, with the exception of the fish family Channichthyidae, as well as the tissues of some invertebrates...
digestion. Gametocytes contained high amounts of gametocyte-specific transcription factors and cell cycle/DNA processing proteins. The gametocytes had low levels of polymorphic surface antigens. Sporozoites contained large amounts of proteins related to invasion, as well as members of the var and rif families.
Metabolism
While all of the metabolic pathways of Plasmodium falciparumPlasmodium falciparum
Plasmodium falciparum is a protozoan parasite, one of the species of Plasmodium that cause malaria in humans. It is transmitted by the female Anopheles mosquito. Malaria caused by this species is the most dangerous form of malaria, with the highest rates of complications and mortality...
have yet to be fully elucidated, the presence and components of many can be predicted through genomic analysis.
Hemoglobin metabolism
During the erythrocytic stage of the parasite's life cycle, it uses intracellular hemoglobinHemoglobin
Hemoglobin is the iron-containing oxygen-transport metalloprotein in the red blood cells of all vertebrates, with the exception of the fish family Channichthyidae, as well as the tissues of some invertebrates...
as a food source. The protein is broken down into peptides, and the heme group is released and detoxified by biocrystallization
Biocrystallization
Biocrystallization is the formation of crystals from organic macromolecules by living organisms. This may be a stress response, a normal part of metabolism such as processes that dispose of waste compounds, or a pathology. Template mediated crystallization is qualitatively different from in vitro...
in the form of hemozoin.
Heme
Heme
A heme or haem is a prosthetic group that consists of an iron atom contained in the center of a large heterocyclic organic ring called a porphyrin. Not all porphyrins contain iron, but a substantial fraction of porphyrin-containing metalloproteins have heme as their prosthetic group; these are...
biosynthesis by the parasite has been reported.
Haemozoin
Ferriprotoporphyrin IX (hematin) competes with NADH for the active site of the enzyme lactate dehydrogenaseLactate dehydrogenase
Lactate dehydrogenase is an enzyme present in a wide variety of organisms, including plants and animals.Lactate dehydrogenases exist in four distinct enzyme classes. Two of them are cytochrome c-dependent enzymes, each acting on either D-lactate or L-lactate...
. This competition may be fatal to the parasite. To detoxify the ferriprotoporphyrin IX the parasite polymerizes hematin to hemozoin which is the characteristic malaria pigment.
Artemisinin
Artemisinin
Artemisinin , also known as Qinghaosu , and its derivatives are a group of drugs that possess the most rapid action of all current drugs against falciparum malaria. Treatments containing an artemisinin derivative are now standard treatment worldwide for falciparum malaria...
an anti malarial agent appears to require haemoglobin digestion for its activity. Inhibition of hemoglobinase activity with cysteine protease inhibitors, knockout of falcipain 2 or deprivation of host cell lysate reduce the activity of the drug against the parasite.
Haemozoin has multiple effects on the hosts immune system including increasing the level of matrix metalloproteinase-9.
Carbohydrate metabolism
During erythrocytic stages, the parasite produces its energy mainly through anaerobic glycolysis, with pyruvate being converted into lactateLactic acid
Lactic acid, also known as milk acid, is a chemical compound that plays a role in various biochemical processes and was first isolated in 1780 by the Swedish chemist Carl Wilhelm Scheele. Lactic acid is a carboxylic acid with the chemical formula C3H6O3...
.
Genes encoding for the TCA cycle enzymes are present in the genome, but it is unclear whether the TCA cycle is used for oxidation of glycolytic products to be used for energy production, or for metabolite intermediate biosynthesis. It has been hypothesized that the main function of the TCA cycle in P. falciparum
Plasmodium falciparum
Plasmodium falciparum is a protozoan parasite, one of the species of Plasmodium that cause malaria in humans. It is transmitted by the female Anopheles mosquito. Malaria caused by this species is the most dangerous form of malaria, with the highest rates of complications and mortality...
is for production of succinyl-CoA, to be used in heme biosynthesis.
Genes for nearly all of the pentose phosphate pathway
Pentose phosphate pathway
The pentose phosphate pathway is a process that generates NADPH and pentoses . There are two distinct phases in the pathway. The first is the oxidative phase, in which NADPH is generated, and the second is the non-oxidative synthesis of 5-carbon sugars...
enzymes have been identified from the genome sequence.
Protein metabolism
It has been hypothesized that the parasite obtains all, or nearly all, of its amino acids by salvaging from the host or through the degradation of hemoglobinHemoglobin
Hemoglobin is the iron-containing oxygen-transport metalloprotein in the red blood cells of all vertebrates, with the exception of the fish family Channichthyidae, as well as the tissues of some invertebrates...
. This is supported by the fact that genomic analysis has found no enzymes necessary for amino acid
Amino acid
Amino acids are molecules containing an amine group, a carboxylic acid group and a side-chain that varies between different amino acids. The key elements of an amino acid are carbon, hydrogen, oxygen, and nitrogen...
biosynthesis, except for glycine-serine, cysteine-alanine, aspartate-asparagine, proline-ornithine, and glutamine-glutamate interconversions
Glutamate-glutamine cycle
In biochemistry, the glutamate-glutamine cycle is a sequence of events by which an adequate supply of the neurotransmitter glutamate is maintained in the central nervous system....
.
Much of this digestion occurs within the digestive vacuole. Multiple enzymes are involved in this process including four distinct plasmepsins (aspartic acid proteases).
Lipid metabolism
PhosphatidylcholinePhosphatidylcholine
Phosphatidylcholines are a class of phospholipids that incorporate choline as a headgroup.They are a major component of biological membranes and can be easily obtained from a variety of readily available sources such as egg yolk or soy beans from which they are mechanically extracted or chemically...
is a major and essential membrane phospholipid in the parasite. Its synthesis occurs via the CDP-choline and the serine decarboxylase phosphoethanolamine methylation pathways. The substrates of these pathways are the host's choline
Choline
Choline is a water-soluble essential nutrient. It is usually grouped within the B-complex vitamins. Choline generally refers to the various quaternary ammonium salts containing the N,N,N-trimethylethanolammonium cation....
, serine
Serine
Serine is an amino acid with the formula HO2CCHCH2OH. It is one of the proteinogenic amino acids. By virtue of the hydroxyl group, serine is classified as a polar amino acid.-Occurrence and biosynthesis:...
and fatty acids. Both pathways share the final two steps catalyzed by two essential enzymes CTP:phosphocholine cytidylyltransferase and choline-phosphate transferase.
β-hydroxyacyl-acyl carrier protein dehydratase catalyzes the third and important reaction of the fatty acid elongation cycle. The P. falciparum gene has been cloned and the crystal structure of the enzyme solved.
Nucleotide metabolism
P. falciparumPlasmodium falciparum
Plasmodium falciparum is a protozoan parasite, one of the species of Plasmodium that cause malaria in humans. It is transmitted by the female Anopheles mosquito. Malaria caused by this species is the most dangerous form of malaria, with the highest rates of complications and mortality...
is unable to biosynthesize purines. Instead, the parasite is able to transport and interconvert host purines.
Conversely, the parasite can produce pyrimidines de novo using glutamine, bicarbonate, and aspartate.
var family
The var genes encode for the P. falciparum erythrocyte membrane protein 1 (PfEMP1) proteins. The genes are found in the subtelomeric regions of the chromosomes. There exist an estimated 59 var genes within the genome.The proteins encoded by the var genes are ultimately transported to the erythrocyte membrane and cause the infected erythrocytes to adhere to host endothelial receptors. Due to transcriptional switching between var genes, antigenic variation occurs which enables immune evasion by the parasite.
The protein PfSir2 associates with promoter regions of silenced genes involved in antigenic variation.
rif family
The rif genes encode for repetitive interspersed family (rifin) proteins. The genes are found in the subtelomeric regions of the chromosomes. There exist an estimated 149 rif genes within the genome.Rifin protein are ultimately transported to the erythrocyte membrane. The function of these proteins is currently unknown.
stevor family
The stevor genes encode for the sub-telomeric variable open reading frame (stevor) proteins. The genes are found in the subtelomeric regions of the chromosomes. There exist an estimated 28 stevor genes within the genome.These proteins appear to affect membrane deformability.
Additional material
Jewett & Sibley (2003) Aldolase forms a bridge between cell surface adhesins and the actin cytoskeleton in apicomplexan parasites. Mol Cell 11(4) 885–94Bergman et al (2003) Myosin A tail domain interacting protein (MTIP) localizes to the inner membrane complex of Plasmodium sporozoites. J Cell Sci 116 (1) 39–49
Baum et al (2005) Invasion by P. falciparum merozoites suggests a hierarchy of molecular interactions. PLoS Pathog 1(4) e37
Bosch et al (2007) The closed MTIP-myosin A-tail complex from the malaria parasite invasion machinery. J Mol Biol 372(1) 77–88
Bosch et al (2007) Aldolase provides an unusual binding site for thrombospondin-related anonymous protein in the invasion machinery of the malaria parasite. Proc Natl Acad Sci USA 104(17) 7015–20
Daher & Soldati-Favre (2009) Mechanisms controlling glideosome function in apicomplexans. Cur Opin Micro 12 (4) 408–414
Sibley (2010) How apicomplexan parasites move in and out of cells. Cur Opin Biotech 21(5) 592–598
External links
- MR4, The NIAID funded Malaria Research and Reference Reagent Resource Center
- PlasmoDB
- Malaria IDC Strain Comparison Database
- Malaria IDC Transcriptome Database
- Malaria Parasite Metabolic Pathways
- ApiCyc