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M13 bacteriophage
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M13 bacteriophage is a ssDNA virus that infects bacteria. The virion is a flexible filament (worm-like chain) about 6 nm in diameter and 900 nm contour length. The genome codes for only 11 genes, so this is one of the simplest organisms known. M13 is 98% identical in DNA sequence to several other filamentous bacteriophage, and this group of phage, including f1, fd and M13, is called the Ff group (they infect bacteria bearing the F fertility factor and they have a filamentous morphology).[1] The viral DNA is encapsidated in a cylindrical protein tube built from several thousand overlapping interdigited copies of the major coat protein, and capped at the two ends with a few copies of minor coat proteins.[2][3][4] The virion infects bacteria by attachment of one of these minor coat proteins to a receptor at the tip of the F pilus of the host bacteria. Unlike most classical bacteriophage, M13 replicates and releases its progeny without killing the host bacteria. M13 is widely used in cloning and other recombinant DNA technology, and has also been used for phage display, directed evolution and nanotechnology applications.[5][6][7] The name "M13" is often used informally as synonymous with other members of the Ff group.
Structure and assembly
[edit]The phage coat is primarily assembled from a 50 amino acid protein called p8, which is encoded by gene 8 in the phage genome. For a wild type M13 particle, it takes approximately 2700 copies of p8 to make the coat about 900 nm long. The coat's dimensions are flexible though and the number of p8 copies adjusts to accommodate the size of the single stranded genome it packages.[8] The phage appear to be limited to approximately twice the natural DNA content. However, deletion of a phage protein (p3) prevents full escape from the host E. coli, and phage that are 10-20X the normal length with several copies of the phage genome can be seen shedding from the E. coli host.
At one end of the filament are five copies of the surface exposed protein (p9) and a more buried companion protein (p7). If p8 forms the shaft of the phage, p9 and p7 form the "blunt" end that is seen in micrographs. These proteins are very small, containing only 33 and 32 amino acids respectively, though some additional residues can be added to the N-terminal portion of each which are then presented on the outside of the coat. At the other end of the phage particle are five copies of the surface exposed (p3) and its less exposed accessory protein (p6). These form the rounded tip of the phage and are the first proteins to interact with the E. coli host during infection. Protein p3 is also the last point of contact with the host as new phage bud from the bacterial surface.
Life cycle
[edit]Below are steps involved with replication of M13 in E. coli.
- Viral (+) strand DNA enters cytoplasm
- Complementary (-) strand is synthesized by bacterial enzymes
- DNA Gyrase, a type II topoisomerase, acts on double-stranded DNA and catalyzes formation of negative supercoils in double-stranded DNA
- Final product is parental replicative form (RF) DNA
- Transcription and translation of the viral genome begins by host resources, including p2.
- A phage protein, p2, nicks the (+) strand in the RF
- 3'-hydroxyl acts as a primer in the creation of new viral strand
- p2 circularizes displaced viral (+) strand DNA
- Pool of progeny double-stranded RF molecules produced
- Negative strand of RF is template of transcription
- mRNAs are translated into the phage proteins
Phage proteins in the cytoplasm are p2, p10 and p5, and they are part of the replication process of DNA. The other phage proteins are synthesized and inserted into the cytoplasmic or outer membranes.
- p5 dimers bind newly synthesized single-stranded DNA and prevent conversion to RF DNA. The timing and attenuation of p5 translation is essential.
- RF DNA synthesis continues and amount of p5 reaches critical concentration
- DNA replication switches to synthesis of single-stranded (+) viral DNA
- p5-DNA structures from about 800 nm long and 8 nm in diameter
- p5-DNA complex is substrate in phage assembly reaction
Applications
[edit]George Smith, among others, showed that fragments of EcoRI endonuclease could be fused in the unique Bam site of f1 filamentous phage and thereby expressed in gene 3 whose protein p3 was externally accessible. M13 does not have this unique Bam site in gene 3. M13 had to be engineered to have accessible insertion sites, making it limited in its flexibility in handling different sized inserts. Because the M13 phage display system[9] allows great flexibility in the location and number of recombinant proteins on the phage, it is a popular tool to construct or serve as a scaffold for nanostructures.[10] For example, the phage can be engineered to have a different protein on each end and along its length. This can be used to assemble structures like gold or cobalt oxide nano-wires for batteries[11] or to pack carbon nanotubes into straight bundles for use in photovoltaics.[12]
See also
[edit]References
[edit]- ^ Rasched, I.; Oberer, E. (1986). "Ff coliphages: structural and functional relationships". Microbiological Reviews. 50 (4): 401–427. ISSN 0146-0749. PMC 373080. PMID 3540571.
{{cite journal}}: CS1 maint: PMC format (link) - ^ Smeal SW, Schmitt MA, Pereira RR, Prasad A, Fisk JD (January 2017). "Simulation of the M13 life cycle I: Assembly of a genetically-structured deterministic chemical kinetic simulation". Virology. 500: 259–274. doi:10.1016/j.virol.2016.08.017. PMID 27644585.
- ^ Rakonjac J, Das B, Derda R (2016). "Editorial: Filamentous Bacteriophage in Bio/Nano/Technology, Bacterial Pathogenesis and Ecology". Frontiers in Microbiology. 7: 2109. doi:10.3389/fmicb.2016.02109. PMC 5179506. PMID 28066406.
{{cite journal}}: CS1 maint: unflagged free DOI (link) - ^ Roux S, Krupovic M, Daly RA, Borges AL, Nayfach S, Schulz F, et al. (November 2019). "Cryptic inoviruses revealed as pervasive in bacteria and archaea across Earth's biomes". Nature Microbiology. 4 (11): 1895–1906. doi:10.1038/s41564-019-0510-x. PMC 6813254. PMID 31332386.
- ^ Khalil AS, Ferrer JM, Brau RR, Kottmann ST, Noren CJ, Lang MJ, Belcher AM (March 2007). "Single M13 bacteriophage tethering and stretching". Proceedings of the National Academy of Sciences of the United States of America. 104 (12): 4892–7. doi:10.1073/pnas.0605727104. PMC 1829235. PMID 17360403.
- ^ Suthiwangcharoen N, Li T, Li K, Thompson P, You S, Wang Q (May 2011). "M13 bacteriophage-polymer nanoassemblies as drug delivery vehicles". Nano Research. 4 (5): 483–93. doi:10.1007/s12274-011-0104-2.
- ^ Esvelt KM, Carlson JC, Liu DR (April 2011). "A system for the continuous directed evolution of biomolecules". Nature. 472 (7344): 499–503. Bibcode:2011Natur.472..499E. doi:10.1038/nature09929. PMC 3084352. PMID 21478873.
- ^ Sattar S, Bennett NJ, Wen WX, Guthrie JM, Blackwell LF, Conway JF, Rakonjac J (2015). "Ff-nano, short functionalized nanorods derived from Ff (f1, fd, or M13) filamentous bacteriophage". Frontiers in Microbiology. 6: 316. doi:10.3389/fmicb.2015.00316. PMC 4403547. PMID 25941520.
{{cite journal}}: CS1 maint: unflagged free DOI (link) - ^ Messing, J. (1996). "Cloning Single-Stranded DNA". Molecular Biotechnology. 5: 39–47.
- ^ Huang Y, Chiang CY, Lee SK, Gao Y, Hu EL, De Yoreo J, Belcher AM (July 2005). "Programmable assembly of nanoarchitectures using genetically engineered viruses". Nano Letters. 5 (7): 1429–34. Bibcode:2005NanoL...5.1429H. doi:10.1021/nl050795d. PMID 16178252.
- ^ Nam KT, Kim DW, Yoo PJ, Chiang CY, Meethong N, Hammond PT, et al. (May 2006). "Virus-enabled synthesis and assembly of nanowires for lithium ion battery electrodes". Science. 312 (5775): 885–8. Bibcode:2006Sci...312..885N. doi:10.1126/science.1122716. PMID 16601154.
- ^ Dang X, Yi H, Ham MH, Qi J, Yun DS, Ladewski R, et al. (April 2011). "Virus-templated self-assembled single-walled carbon nanotubes for highly efficient electron collection in photovoltaic devices". Nature Nanotechnology. 6 (6): 377–84. Bibcode:2011NatNa...6..377D. doi:10.1038/nnano.2011.50. PMID 21516089.
Further reading
[edit]- Barbas, Carlos F; Burton, Dennis R; Silverman, Gregg J (October 2004). Phage Display: A Laboratory Manual (1st ed.). Cold Spring Harbor Laboratory Press. ISBN 978-0-87969-740-2.
{{cite book}}: Unknown parameter|name-list-format=ignored (|name-list-style=suggested) (help) - Messing J (1993). "M13 Cloning Vehicles" (PDF). In Griffin H.G., Griffin A.M. (eds.). DNA Sequencing Protocols. Methods in Molecular Biology™. Methods in Molecular Biology. Vol. 23. Humana Press. pp. 9–22. doi:10.1385/0-89603-248-5:9. ISBN 0-89603-248-5. PMID 8220775. Archived from the original on 2012-02-19.