Proteomics - AbsoluteAstronomy.com

Proteomics is the large-scale study of 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, particularly their structures

Protein structure

Proteins are an important class of biological macromolecules present in all organisms. Proteins are polymers of amino acids. Classified by their physical size, proteins are nanoparticles . Each protein polymer – also known as a polypeptide – consists of a sequence formed from 20 possible L-α-amino...

and functions

Functional genomics

Functional genomics is a field of molecular biology that attempts to make use of the vast wealth of data produced by genomic projects to describe gene functions and interactions...

. Proteins are vital parts of living organisms, as they are the main components of the physiological metabolic pathways of cells. The term "proteomics" was first coined in 1997 to make an analogy with genomics

Genomics

Genomics is a discipline in genetics concerning the study of the genomes of organisms. The field includes intensive efforts to determine the entire DNA sequence of organisms and fine-scale genetic mapping efforts. The field also includes studies of intragenomic phenomena such as heterosis,...

, the study of the gene

Gene

A gene is a molecular unit of heredity of a living organism. It is a name given to some stretches of DNA and RNA that code for a type of protein or for an RNA chain that has a function in the organism. Living beings depend on genes, as they specify all proteins and functional RNA chains...

s. The word "proteome" is a blend

Blend

In linguistics, a blend is a word formed from parts of two or more other words. These parts are sometimes, but not always, morphemes.-Linguistics:...

of "protein" and "genome", and was coined by Marc Wilkins

Marc Wilkins (geneticist)

Marc R. Wilkins is an Australian scientist who is credited with the concept of the proteome, and is a Professor in the School of Biotechnology and Biomolecular Sciences at the University of New South Wales, Sydney, Australia....

in 1994 while working on the concept as a PhD student. 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 the entire complement of proteins, including the modifications made to a particular set of proteins, produced by an organism or system. This will vary with time and distinct requirements, or stresses, that a cell or organism undergoes.

Complexity of the problem

After genomics, proteomics is considered the next step in the study of biological systems. It is much more complicated than genomics mostly because while an organism's 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....

is more or less constant, the proteome

Proteome

differs from cell to cell and from time to time. This is because distinct genes are expressed in distinct cell types. This means that even the basic set of proteins which are produced in a cell needs to be determined.

In the past this was done by mRNA analysis, but this was found not to correlate with protein content. It is now known that mRNA is not always translated into protein, and the amount of protein produced for a given amount of mRNA depends on the gene it is transcribed from and on the current physiological state of the cell. Proteomics confirms the presence of the protein and provides a direct measure of the quantity present.

Post-translational modifications

Not only does the translation from mRNA cause differences, many proteins are also subjected to a wide variety of chemical modifications after translation. Many of these post-translational modifications are critical to the protein's function.

Phosphorylation

One such modification is phosphorylation

Phosphorylation

Phosphorylation is the addition of a phosphate group to a protein or other organic molecule. Phosphorylation activates or deactivates many protein enzymes....

, which happens to many enzymes and structural proteins in the process of cell signaling

Cell signaling

Cell signaling is part of a complex system of communication that governs basic cellular activities and coordinates cell actions. The ability of cells to perceive and correctly respond to their microenvironment is the basis of development, tissue repair, and immunity as well as normal tissue...

. The addition of a phosphate to particular amino acids—most commonly 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 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...

mediated by serine/threonine kinase

Kinase

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, or more rarely tyrosine

Tyrosine

Tyrosine or 4-hydroxyphenylalanine, is one of the 22 amino acids that are used by cells to synthesize proteins. Its codons are UAC and UAU. It is a non-essential amino acid with a polar side group...

mediated by tyrosine kinases—causes a protein to become a target for binding or interacting with a distinct set of other proteins that recognize the phosphorylated domain.

Because protein phosphorylation is one of the most-studied protein modifications, many "proteomic" efforts are geared to determining the set of phosphorylated proteins in a particular cell or tissue-type under particular circumstances. This alerts the scientist to the signaling pathways that may be active in that instance.

Ubiquitination

Ubiquitin

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...

is a small protein that can be affixed to certain protein substrates by enzymes called E3 ubiquitin ligases. Determining which proteins are poly-ubiquitinated can be helpful in understanding how protein pathways are regulated. This is therefore an additional legitimate "proteomic" study. Similarly, once it is determined which substrates are ubiquitinated by each ligase, determining the set of ligases expressed in a particular cell type will be helpful.

Additional modifications

Listing all the protein modifications that might be studied in a "Proteomics" project would require a discussion of most of biochemistry

Biochemistry

Biochemistry, sometimes called biological chemistry, is the study of chemical processes in living organisms, including, but not limited to, living matter. Biochemistry governs all living organisms and living processes...

; therefore, a short list will serve here to illustrate the complexity of the problem.
In addition to phosphorylation

Phosphorylation

Phosphorylation is the addition of a phosphate group to a protein or other organic molecule. Phosphorylation activates or deactivates many protein enzymes....

and ubiquitination, proteins can be subjected to (among others) methylation

Methylation

In the chemical sciences, methylation denotes the addition of a methyl group to a substrate or the substitution of an atom or group by a methyl group. Methylation is a form of alkylation with, to be specific, a methyl group, rather than a larger carbon chain, replacing a hydrogen atom...

, acetylation

Acetylation

Acetylation describes a reaction that introduces an acetyl functional group into a chemical compound...

, glycosylation

Glycosylation

Glycosylation is the reaction in which a carbohydrate, i.e. a glycosyl donor, is attached to a hydroxyl or other functional group of another molecule . In biology glycosylation refers to the enzymatic process that attaches glycans to proteins, lipids, or other organic molecules...

, oxidation and nitrosylation

Nitrosylation

Nitrosylation is a protein modification in which a nitrosyl group is post-translationally added to a protein.There is a range of enzymes that produce nitric oxide, and the frequent consequence of this production is nitrosylation....

. Some proteins undergo ALL of these modifications, often in time-dependent combinations, aptly illustrating the potential complexity one has to deal with when studying protein structure and function.

Distinct proteins are made under distinct settings

Even if one is studying a particular cell type, that cell may make different sets of proteins at different times, or under different conditions. Furthermore, as mentioned, any one protein can undergo a wide range of post-translational modifications.

Therefore a "proteomics" study can become quite complex very quickly, even if the object of the study is very restricted. In more ambitious settings, such as when a biomarker for a tumor is sought – when the proteomics scientist is obliged to study sera samples from multiple cancer patients – the amount of complexity that must be dealt with is as great as in any modern biological project.

Limitations to genomic study

Scientists are very interested in proteomics because it gives a much better understanding of an organism than genomics. First, the level of transcription of a gene gives only a rough estimate of its level of expression into a protein. An mRNA produced in abundance may be degraded rapidly or translated inefficiently, resulting in a small amount of protein. Second, as mentioned above many proteins experience post-translational modifications that profoundly affect their activities; for example some proteins are not active until they become phosphorylated. Methods such as phosphoproteomics

Phosphoproteomics

Phosphoproteomics is a branch of proteomics that identifies, catalogs, and characterizes proteins containing a phosphate group as a post-translational modification. Phosphorylation is a key reversible modification that regulates protein function, subcellular localization, complex formation,...

and glycoproteomics

Glycoproteomics

Glycoproteomics is a branch of proteomics that identifies, catalogs, and characterizes proteins containing carbohydrates as a post-translational modification....

are used to study post-translational modifications. Third, many transcripts give rise to more than one protein, through alternative splicing

Alternative splicing

Alternative splicing is a process by which the exons of the RNA produced by transcription of a gene are reconnected in multiple ways during RNA splicing...

or alternative post-translational modifications. Fourth, many proteins form complexes with other proteins or RNA molecules, and only function in the presence of these other molecules. Finally, protein degradation rate plays an important role in protein content.

Determining proteins which are post-translationally modified

One way in which a particular protein can be studied is to develop an antibody

Antibody

An antibody, also known as an immunoglobulin, is a large Y-shaped protein used by the immune system to identify and neutralize foreign objects such as bacteria and viruses. The antibody recognizes a unique part of the foreign target, termed an antigen...

which is specific to that modification. For example, there are antibodies which only recognize certain proteins when they are tyrosine-phosphorylated, known as phospho-specific antibodies; also, there are antibodies specific to other modifications. These can be used to determine the set of proteins that have undergone the modification of interest.

For sugar

Sugar

Sugar is a class of edible crystalline carbohydrates, mainly sucrose, lactose, and fructose, characterized by a sweet flavor.Sucrose in its refined form primarily comes from sugar cane and sugar beet...

modifications, such as glycosylation

Glycosylation

of proteins, certain lectins have been discovered which bind sugars. These too can be used.

A more common way to determine post-translational modification of interest is to subject a complex mixture of proteins to electrophoresis in "two-dimensions", which simply means that the proteins are electrophoresed first in one direction, and then in another, which allows small differences in a protein to be visualized by separating a modified protein from its unmodified form. This methodology is known as "two-dimensional gel electrophoresis

Two-dimensional gel electrophoresis

Two-dimensional gel electrophoresis, abbreviated as 2-DE or 2-D electrophoresis, is a form of gel electrophoresis commonly used to analyze proteins...

".

Recently, another approach has been developed called PROTOMAP

Protomap (proteomics)

PROTOMAP is a recently developed proteomic technology for identifying changes to proteins that manifest in altered migration by one-dimensional SDS-PAGE...

which combines SDS-PAGE

SDS-PAGE

SDS-PAGE, sodium dodecyl sulfate polyacrylamide gel electrophoresis, describes a collection of related techniques widely used in biochemistry, forensics, genetics and molecular biology to separate proteins according to their electrophoretic mobility...

with shotgun proteomics

Shotgun proteomics

Shotgun proteomics is a method of identifying proteins in complex mixtures using a combination of high performance liquid chromatography combined with mass spectrometry. The name is derived from shotgun sequencing of DNA which is itself named by analogy with the rapidly-expanding, quasi-random...

to enable detection of changes in gel-migration such as those caused by proteolysis

Proteolysis

Proteolysis is the directed degradation of proteins by cellular enzymes called proteases or by intramolecular digestion.-Purposes:Proteolysis is used by the cell for several purposes...

or post translational modification.

Determining the existence of proteins in complex mixtures

Classically, antibodies to particular proteins or to their modified forms have been used in biochemistry

Biochemistry

and cell biology

Cell biology

Cell biology is a scientific discipline that studies cells – their physiological properties, their structure, the organelles they contain, interactions with their environment, their life cycle, division and death. This is done both on a microscopic and molecular level...

studies. These are among the most common tools used by practicing biologists today.

For more quantitative determinations of protein amounts, techniques such as ELISA

ELISA

Enzyme-linked immunosorbent assay , is a popular format of a "wet-lab" type analytic biochemistry assay that uses one sub-type of heterogeneous, solid-phase enzyme immunoassay to detect the presence of a substance in a liquid sample."Wet lab" analytic biochemistry assays involves detection of an...

s can be used.

For proteomic study, more recent techniques such as matrix-assisted laser desorption/ionization (MALDI)

Matrix-assisted laser desorption/ionization

Matrix-assisted laser desorption/ionization is a soft ionization technique used in mass spectrometry, allowing the analysis of biomolecules and large organic molecules , which tend to be fragile and fragment when ionized by more conventional ionization methods...

have been employed for rapid determination of proteins in particular mixtures and increasingly electrospray ionization (ESI)

Electrospray ionization

Electrospray ionization is a technique used in mass spectrometry to produce ions. It is especially useful in producing ions from macromolecules because it overcomes the propensity of these molecules to fragment when ionized...

Computational methods in studying protein biomarkers

Computational predictive models have shown that extensive and diverse feto-maternal protein trafficking occurs during pregnancy and can be readily detected non-invasively in maternal whole blood. This computational approach circumvented a major limitation, the abundance of maternal proteins interfering with the detection of fetal protein

Fetal protein

Fetal proteins are simply proteins present at highest levels at the fetus stage. Often related proteins assume similar roles after birth or in the embryo, in which case the fetal varieties are called fetal isoforms...

s, to fetal proteomic analysis of maternal blood. Computational models can use fetal gene transcripts previously identified in maternal whole blood

Whole blood

Whole blood is a term used in transfusion medicine for human blood from a standard blood donation. The blood is typically combined with an anticoagulant during the collection process, but is generally otherwise unprocessed...

to create a comprehensive proteomic network of the term neonate. Such work shows that the fetal proteins detected in pregnant woman’s blood originate from a diverse group of tissues and organs from the developing fetus. The proteomic networks contain many biomarkers

Biomarker (medicine)

In medicine, a biomarker is a term often used to refer to a protein measured in blood whose concentration reflects the severity or presence of some disease state...

that are proxies for development and illustrate the potential clinical application of this technology as a way to monitor normal and abnormal fetal development.

An information theoretic framework has also been introduced for biomarker

Biomarker (medicine)

In medicine, a biomarker is a term often used to refer to a protein measured in blood whose concentration reflects the severity or presence of some disease state...

discovery, integrating biofluid and tissue information. This new approach takes advantage of functional synergy between certain biofluids and tissues with the potential for clinically significant findings not possible if tissues and biofluids were considered individually. By conceptualizing tissue-biofluid as information channels, significant biofluid proxies can be identified and then used for guided development of clinical diagnostics. Candidate biomarkers are then predicted based on information transfer criteria across the tissue-biofluid channels. Significant biofluid-tissue relationships can be used to prioritize clinical validation of biomarkers.

Establishing protein–protein interactions

Most proteins function in collaboration with other proteins, and one goal of proteomics is to identify which proteins interact. This is especially useful in determining potential partners in cell signaling

Cell signaling

cascades.

Several methods are available to probe protein–protein interactions. The traditional method is yeast two-hybrid analysis

Two-hybrid screening

Two-hybrid screening is a molecular biology technique used to discover protein–protein interactions and protein–DNA interactions by testing for physical interactions between two proteins or a single protein and a DNA molecule, respectively.The premise behind the test is the activation of...

. New methods include protein microarray

Protein microarray

A protein microarray, sometimes referred to as a protein binding microarray,provides a multiplex approach to identify protein–protein interactions, to identify the substrates of protein kinases, to identify transcription factor protein-activation, or to identify the targets of biologically active...

s, immunoaffinity chromatography followed by mass spectrometry

Mass spectrometry

Mass spectrometry is an analytical technique that measures the mass-to-charge ratio of charged particles.It is used for determining masses of particles, for determining the elemental composition of a sample or molecule, and for elucidating the chemical structures of molecules, such as peptides and...

, dual polarisation interferometry

Dual Polarisation Interferometry

Dual polarization interferometry is an analytical technique that can probe molecular scale layers adsorbed to the surface of a waveguide by using the evanescent wave of a laser beam confined to the waveguide...

, Microscale Thermophoresis

Microscale Thermophoresis

Microscale Thermophoresis is a technology for the analysis of biomolecules. Microscale Thermophoresis is the directed movement of particles in a microscopic temperature gradient...

and experimental methods such as phage display and computational methods

Practical applications of proteomics

One of the most promising developments to come from the study of human genes and proteins has been the identification of potential new drugs for the treatment of disease. This relies on genome and proteome information to identify proteins associated with a disease, which computer software can then use as targets for new drugs. For example, if a certain protein is implicated in a disease, its 3D structure provides the information to design drugs to interfere with the action of the protein. A molecule that fits the active site of an enzyme, but cannot be released by the enzyme, will inactivate the enzyme. This is the basis of new drug-discovery tools, which aim to find new drugs to inactivate proteins involved in disease. As genetic differences among individuals are found, researchers expect to use these techniques to develop personalized drugs that are more effective for the individual.

Biomarkers

The FDA defines a biomarker as, “A characteristic that is objectively measured and evaluated as an indicator of normal biologic processes, pathogenic processes, or pharmacologic responses to a therapeutic intervention”.

Understanding the proteome, the structure and function of each protein and the complexities of protein–protein interactions will be critical for developing the most effective diagnostic techniques and disease treatments in the future.

An interesting use of proteomics is using specific protein biomarkers to diagnose disease. A number of techniques allow to test for proteins produced during a particular disease, which helps to diagnose the disease quickly. Techniques include western blot

Western blot

The western blot is a widely used analytical technique used to detect specific proteins in the given sample of tissue homogenate or extract. It uses gel electrophoresis to separate native proteins by 3-D structure or denatured proteins by the length of the polypeptide...

, immunohistochemical staining, enzyme linked immunosorbent assay (ELISA) or mass spectrometry

Mass spectrometry

. Secretomics

Secretomics

Secretomics is a subset of proteomics in which all of the secreted proteins of a cell, tissue, or organism are analyzed. Secreted proteins are involved in a variety of physiological processes, including cell signaling and matrix remodeling, but are also integral to invasion and metastasis of...

, a subfield of proteomics that studies secreted proteins

Secretory protein

A secretory protein is any protein, whether it be endocrine or exocrine, which is secreted by a cell. Secretory proteins include many hormones, enzymes, toxins, and antimicrobial peptides.Secretory proteins are synthesized in endoplasmic reticulum....

and secretion pathways using proteomic approaches, has recently emerged as an important tool for the discovery of biomarkers of disease.

Proteogenomics

In what is now commonly referred to as proteogenomics

Proteogenomics

Proteogenomics is an emerging field of biological research at the intersection of proteomics and genomics. While this intersection is large and can be defined in multiple ways, the term proteogenomics commonly refers to studies that use proteomic information, often derived from mass spectrometry,...

, proteomic technologies such as mass spectrometry

Mass spectrometry

are used for improving gene annotations. Parallel analysis of the genome and the proteome facilitates discovery of post-translational modifications and proteolytic events , especially when comparing multiple species (comparative proteogenomics).

Current research methodologies

Fluorescence two-dimensional differential gel electrophoresis (2-D DIGE) can be used to quantify variation in the 2-D DIGE process and establish statistically valid thresholds for assigning quantitative changes between samples.

Comparative proteomic analysis can reveal the role of proteins in complex biological systems, including reproduction. For example, treatment with the insecticide triazophos causes an increase in the content of brown planthopper (Nilaparvata lugens (Stål)) male accessory gland proteins (Acps) that can be transferred to females via mating, causing an increase in fecundity (i.e. birth rate) of females. To identify changes in the types of accessory gland proteins (Acps) and reproductive proteins that mated female planthoppers received from male planthoppers, researchers conducted a comparative proteomic analysis of mated N. lugens females. The results indicated that these proteins participate in the reproductive process of N. lugens adult females and males.

Proteome analysis of Arabidopsis peroxisomes has been established as the major unbiased approach for identifying new peroxisomal proteins on a large scale.

There are many approaches to characterizing the human proteome, which is estimated to contain between 20,000 and 25,000 non-redundant proteins. The number of unique protein species likely increase by between 50,000 and 500,000 due to RNA splicing and proteolysis events, and when post-translational modification are also considered, the total number of unique human proteins is estimated to range in the low millions.

In addition, first promising attempts to decipher the proteome of animal tumors have recently been reported.

External links

Easily accessible Proteomics information

The source of this article is wikipedia, the free encyclopedia. The text of this article is licensed under the GFDL.

Complexity of the problem

Post-translational modifications

Phosphorylation

Ubiquitination

Additional modifications

Distinct proteins are made under distinct settings

Limitations to genomic study

Determining proteins which are post-translationally modified

Determining the existence of proteins in complex mixtures

Computational methods in studying protein biomarkers

Establishing protein–protein interactions

Practical applications of proteomics

Biomarkers

Proteogenomics

Current research methodologies

See also

Protein databases

External links