Phi-X174 phage
Encyclopedia
The phi X 174 bacteriophage
was the first DNA-based genome
to be sequenced. This work was completed by Fred Sanger and his team in 1977. In 1962, Walter Fiers
had already demonstrated the physical, covalently closed circularity of phi X 174 DNA.
In 2003, it was reported that the whole genome of ΦX174 had been assembled synthetically from scratch. ΦX174 has also been successfully assembled in vitro
has a [+] circular single-stranded DNA
genome of 5386 nucleotide
s encoding 11 protein
s. Of these 11 genes, only 8 are essential to viral morphogenesis. The GC-content
is 44% and 95% of nucleotides belong to coding genes.
Table from ΦX174 et al. the Microviridae by B.A. Fane et al.
Infection begins when H protein (or the DNA Pilot Protein) pilots the viral genome through the bacterial membrane of E.coli bacteria (Jazwinski et al. 1975) most likely via a predicted N-terminal transmembrane domain helix (Tusnady and Simon, 2001). However, it has become apparent that H protein is a multifunctional protein (Cherwa, Young and Fane, 2011). This is the only viral capsid protein of ΦX174 to lack a crystal structure for a couple of reasons. It has low aromatic content and high glycine content, making the protein structure very flexible and in addition, individual hydrogen atoms (the R group for glycines) are difficult to detect in protein crystallography. Additionally, H protein induces lysis
of the bacterial host at high concentrations as the predicted N-terminal transmembrane helix easily pokes holes through the bacterial wall. By bioinformatics, this protein contains four predicted coiled-coil domains which has a significant homology to known transcription factors. Additionally, it was determined by Ruboyianes et al. (2009) that de novo H protein was required for optimal synthesis of other viral proteins. Interestingly, mutations in H protein that prevent viral incorporation, can be overcome when excess amounts of Protein B, the internal scaffolding protein, are supplied.
The DNA is ejected through a hydrophilic channel at the 5-fold vertex (McKenna et al. 1992). It is understood that H protein resides in this area but experimental evidence has not verified its exact location. Once inside the host bacterium, replication of the [+] ssDNA genome proceeds via a rolling circle mechanism. As D protein is the most abundant gene transcript, it is the most protein in the viral procaspid. Similarly, gene transcripts for F, J, and G are more abundant than for H as the stoichiometry for these structures proteins is 5:5:5:1.
Bacteriophage
A bacteriophage is any one of a number of viruses that infect bacteria. They do this by injecting genetic material, which they carry enclosed in an outer protein capsid...
was the first DNA-based 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....
to be sequenced. This work was completed by Fred Sanger and his team in 1977. In 1962, Walter Fiers
Walter Fiers
Walter Fiers is a Belgian molecular biologist.He obtained a degree of Engineer for Chemistry and Agricultural Industries at the University of Ghent in 1954, and started his research career as an enzymologist in the laboratory of Laurent Vandendriessche in Ghent. In 1956-57, he worked with Heinz...
had already demonstrated the physical, covalently closed circularity of phi X 174 DNA.
In 2003, it was reported that the whole genome of ΦX174 had been assembled synthetically from scratch. ΦX174 has also been successfully assembled in vitro
Virology
This bacteriophageBacteriophage
A bacteriophage is any one of a number of viruses that infect bacteria. They do this by injecting genetic material, which they carry enclosed in an outer protein capsid...
has a [+] circular single-stranded 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...
genome of 5386 nucleotide
Nucleotide
Nucleotides are molecules that, when joined together, make up the structural units of RNA and DNA. In addition, nucleotides participate in cellular signaling , and are incorporated into important cofactors of enzymatic reactions...
s encoding 11 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. Of these 11 genes, only 8 are essential to viral morphogenesis. The GC-content
GC-content
In molecular biology and genetics, GC-content is the percentage of nitrogenous bases on a DNA molecule that are either guanine or cytosine . This may refer to a specific fragment of DNA or RNA, or that of the whole genome...
is 44% and 95% of nucleotides belong to coding genes.
| Protein | Function |
|---|---|
| A | Stage II and stage III DNA replication |
| A* | An unessential protein for viral propagation. It may play a role in the inhibition of host cell DNA replication and superinfection exclusion |
| B | Internal scaffolding protein, required for capsid morphogenesis and the assembly of early morphogenetic intermediates. Sixty copies present in the procapsid |
| C | Facilitates the switch from stage II to stage III DNA replication. Required for stage III DNA synthesis |
| D | External scaffolding protein, required for procapsid morphogenesis. Two hundred and forty copies present in the procapsid. |
| E | Host cell lysis |
| F | Major coat protein. Sixty copies present in the virion and procapsid |
| G | Major spike protein. Sixty copies present in the virion and procapsid |
| H | DNA pilot protein need for DNA injection, also called the minor spike protein. Twelve copies in the procapsid and virion |
| J | DNA binding protein, needed for DNA packaging. Sixty copies present in the virion |
| K | An unessential protein for viral propagation. It may play a role optimizing burst sizes in various Hosts |
Table from ΦX174 et al. the Microviridae by B.A. Fane et al.
Infection begins when H protein (or the DNA Pilot Protein) pilots the viral genome through the bacterial membrane of E.coli bacteria (Jazwinski et al. 1975) most likely via a predicted N-terminal transmembrane domain helix (Tusnady and Simon, 2001). However, it has become apparent that H protein is a multifunctional protein (Cherwa, Young and Fane, 2011). This is the only viral capsid protein of ΦX174 to lack a crystal structure for a couple of reasons. It has low aromatic content and high glycine content, making the protein structure very flexible and in addition, individual hydrogen atoms (the R group for glycines) are difficult to detect in protein crystallography. Additionally, H protein induces lysis
Lysis
Lysis refers to the breaking down of a cell, often by viral, enzymic, or osmotic mechanisms that compromise its integrity. A fluid containing the contents of lysed cells is called a "lysate"....
of the bacterial host at high concentrations as the predicted N-terminal transmembrane helix easily pokes holes through the bacterial wall. By bioinformatics, this protein contains four predicted coiled-coil domains which has a significant homology to known transcription factors. Additionally, it was determined by Ruboyianes et al. (2009) that de novo H protein was required for optimal synthesis of other viral proteins. Interestingly, mutations in H protein that prevent viral incorporation, can be overcome when excess amounts of Protein B, the internal scaffolding protein, are supplied.
The DNA is ejected through a hydrophilic channel at the 5-fold vertex (McKenna et al. 1992). It is understood that H protein resides in this area but experimental evidence has not verified its exact location. Once inside the host bacterium, replication of the [+] ssDNA genome proceeds via a rolling circle mechanism. As D protein is the most abundant gene transcript, it is the most protein in the viral procaspid. Similarly, gene transcripts for F, J, and G are more abundant than for H as the stoichiometry for these structures proteins is 5:5:5:1.