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Virus structure and mechanics information and references
Virus structure and mechanics information and references
General information about viruses
P22 virion
Other virions
DNA packaging molecular motors
General information about viruses
- The Wikipedia article on Virus is particularly good on a range of topics.
With respect to virus origins:
Viruses are found wherever there is life and have probably existed since living cells first evolved.[27] The origin of viruses is unclear because they do not form fossils, so molecular techniques have been the most useful means of investigating how they arose.[28] These techniques rely on the availability of ancient viral DNA or RNA, but, unfortunately, most of the viruses that have been preserved and stored in laboratories are less than 90 years old.[29][30] There are three main theories of the origins of viruses:[31][32]
- Regressive theory: Viruses may have once been small cells that parasitised larger cells. Over time, genes not required by their parasitism were lost. The bacteria rickettsia and chlamydia are living cells that, like viruses, can reproduce only inside host cells. They lend credence to this theory, as their dependence on parasitism is likely to have caused the loss of genes that enabled them to survive outside a cell. This is also called the degeneracy theory.[33][34]
- Cellular origin theory (sometimes called the vagrancy theory):[33][35] Some viruses may have evolved from bits of DNA or RNA that "escaped" from the genes of a larger organism. The escaped DNA could have come from plasmids—pieces of naked DNA that can move between cells or transposons. These are molecules of DNA that replicate and move around to different positions within the genes of the cell.[36] Once called "jumping genes", these are examples of mobile genetic elements and could be the origin of some viruses. Transposons were discovered in maize by Barbara McClintock in 1950.[37]
- Coevolution theory: Viruses may have evolved from complex molecules of protein and nucleic acid at the same time as cells first appeared on earth and would have been dependent on cellular life for many millions of years. Viroids are molecules of RNA that are not classified as viruses because they lack a protein coat. However, they have characteristics which are common to several viruses and are often called subviral agents.[38] Viroids are important pathogens of plants.[39] They do not code for proteins but interact with the host cell and use the host machinery for their replication.[40] The hepatitis delta virus of humans has an RNA genome similar to viroids but has protein coat derived from hepatitis B virus and cannot produce one of its own. It is therefore a defective virus and cannot replicate without the help of hepatitis B virus.[41] These viruses that are dependent on other virus species are called satellites and may represent evolutionary intermediates of viroids and viruses.[42][43] Prions are infectious protein molecules that do not contain DNA or RNA.[44] They cause an infection in sheep called scrapie and cattle bovine spongiform encephalopathy ("mad cow" disease). In humans they cause kuru and Creutzfeld-Jacob disease.[45] They are able to replicate because some proteins can exist in two different shapes and the prion changes the normal shape of a host protein into the prion shape. This starts a chain reaction where each prion protein converts many host proteins into more prions, and these new prions then go on to convert even more protein into prions. Although they are fundamentally different from viruses and viroids, there discovery gives credence to theory that viruses could have evolved from self-replicating molecules.[46]
Computer analysis of viral and host DNA sequences is giving a better understanding of the evolutionary relationships between different viruses and may help identify the ancestors of modern viruses. To date, such analyses have not helped to decide on which of the theories are correct. However, it seems unlikely that all currently known viruses have a common ancestor and viruses have probably arisen numerous times in the past by one or more mechanisms.[47]
Opinions differ on whether viruses are a form of life, or organic structures that interact with living organisms. They have been described as "organisms at the edge of life",[48] since they resemble organisms in that they possess genes and evolve by natural selection,[49] and reproduce by creating multiple copies of themselves through self-assembly. However, although they have genes, they do not have a cellular structure, which is often seen as the basic unit of life. Additionally, viruses do not have their own metabolism, and require a host cell to make new products. They therefore cannot reproduce outside a host cell (though bacterial species such as rickettsia and chlamydia are considered living organisms despite the same limitation). Accepted forms of life use cell division to reproduce, whereas viruses spontaneously assemble within cells, which is analogous to the autonomous growth of crystals. Virus self-assembly within host cells has implications for the study of the origin of life,[50] as it lends further credence to the hypothesis that life could have started as self-assembling organic molecules.[51]
With respect to structure:
Viruses display a wide diversity of shapes and sizes, called morphologies. Viruses are about 100 times smaller than bacteria. Most viruses which have been studied have a diameter between 10 and 300 nanometres. Some filoviruses have a total length of up to 1400 nm, however their diameters are only about 80 nm.[3] Most viruses are unable to be seen with a light microscope so scanning and transmission electron microscopes are used to visualise virus particles.[52] To increase the contrast between viruses and the background, electron-dense "stains" are used. These are solutions of salts of heavy metals such as tungsten, that scatter the electrons from regions covered with the stain. When virus particles are coated with stain (positive staining), fine detail is obscured. Negative staining overcomes this problem by staining the background only.[53]
A complete virus particle, known as a virion, consists of nucleic acid surrounded by a protective coat of protein called a capsid. These are formed from identical protein subunits called capsomers.[54] Viruses can have a lipid "envelope" derived from the host cell membrane. The capsid is made from proteins encoded by the viral genome and its shape serves as the basis for morphological distinction.[55][56] Virally coded protein subunits will self-assemble to form a capsid, generally requiring the presence of the virus genome. However, complex viruses code for proteins which assist in the construction of their capsid. Proteins associated with nucleic acid are known as nucleoproteins, and the association of viral capsid proteins with viral nucleic acid is called a nucleocapsid. In general, there are four main morphological virus types:
Helical capsids are composed of a single type of capsomer stacked around a central axis to form a helical structure which may have a central cavity, or hollow tube. This arrangement results in rod-shaped or filamentous virions: these can be short and highly rigid, or long and very flexible. The genetic material, generally single-stranded RNA, but ssDNA in some cases, is bound into the protein helix, by interactions between the negatively-charged nucleic acid and positive charges on the protein. Overall, the length of a helical capsid is related to the length of the nucleic acid contained within it and the diameter is dependent on the size and arrangement of capsomers. The well-studied Tobacco mosaic virus is an example of a helical virus.[57]
Most animal viruses are icosahedral or near-spherical with icosahedral symmetry. A regular icosahedron is the optimum way of forming a closed shell from identical sub-units. The minimum number of identical capsomers required is twelve, each composed of five identical sub-units. Many viruses, such as rotavirus, have more than twelve capsomers and appear spherical but they retain this symmetry. Capsomers at the apices are surrounded by five other capsomers and are called pentons. Capsomers on the triangular faces are surround by six others and are call hexons.[58]
Some species of viruses envelope themselves in a modified form of one of the cell membranes, either the outer membrane surrounding an infected host cell, or internal membranes such as nuclear membrane or endoplasmic reticulum, thus gaining an outer lipid bilayer known as a viral envelope. This membrane is studded with proteins coded for by the viral genome and host genome; the lipid membrane itself and any carbohydrates present are entirely host-coded. The influenza virus and HIV use this strategy. Most enveloped viruses are dependent on the envelope for their infectivity.[59]
These viruses possess a capsid which is neither purely helical, nor purely icosahedral, and which may possess extra structures such as protein tails or a complex outer wall. Some bacteriophages have a complex structure consisting of an icosahedral head bound to a helical tail which may have a hexagonal base plate with protruding protein tail fibres.
The poxviruses are large, complex viruses which have an unusual morphology. The viral genome is associated with proteins within a central disk structure known as a nucleoid. The nucleoid is surrounded by a membrane and two lateral bodies of unknown function. The virus has an outer envelope with a thick layer of protein studded over its surface. The whole particle is slightly pleiomorphic, ranging from ovoid to brick shape.[60] Mimivirus is the largest known virus, with a capsid diameter of 400 nm. Protein filaments measuring 100 nm project from the surface. The capsid appears hexagonal under an electron microscope, therefore the capsid is probably icosahedral.[61]
- From Wikipedia Michael Rossmann:
In 1985, he published his team's mapping, using X-ray crystallography, of a human common cold virus in the journal Nature[4]. The breakthrough nature of this result was such that the National Science Foundation, which provided partial funding for the research, saw fit to organize a press conference, and the news travelled in the general press.[5]
Virus and virion data sources
- Macromolecular Structure Database Group, part of European Bioinformatics Institute's Protein Data Bank Europe (PDBe). Also see the EM Data Bank, which provides three-dimensional density maps for non-viruses.
- VIPERdb:
- This web site describes various icosahedral virus capsid structures in the Protein Data Bank (PDB) in terms of their complete capsids, detailed structural and computational analysis.
- The protomeric (asymmetric unit) coordinates were transformed and stored in a single icosahedral convention (2(Z)-3-5-(X)2). Coordinates (sometimes) were further processed by rearranging the nucleic acid residues, hetero atoms, and water molecules (if present) at the end of the protein chains for the ease of further analysis.
- Structures are classified in terms of their quasi-symmetry (e.g. T=3, T=4, T=7) and pseudo-symmetry (e.g., P=3).
- Tools are being developed to study the capsids in terms of structural, energetic and assembly aspects . The derived results are made available throught this website.
P22 overviews
P22 is a tailed bacteriophage which are in the order Caudovirales .
- Information about bacteriophage P22
by Sherwood Casjens (sherwood.casjens@path.utah.edu)
Electron micrograph of bacteriophage P22
History, Fame, & Fortune
Morphology & classification
Relatives
Hosts & cultivation of this phage
Genomics
Assembly pathway
References
These are mostly recent references to give the reader access to up-to-date literature on the subject. Original references can be found within them.
1. Zinder, N., and Lederberg, J. (1952) Genetic exchange in Salmonella. J. Bacteriol. 64: 679-699.
2. Masters, M. (1996). Generalized transduction. in Escherichia coli and Salmonella typhimurium (Neidhardt, F., Ed.-in-chief), ASM press, Washington, D.C., pp2412-2441
3. Ebel-Tsipsis, J., Botstein, D., and Fox, M. (1972) Generalized transduction by phage P22 in Salmonella typhimurium. J. Mol. Biol. 71: 433-448.
4. Lobban, P., and Kaiser, A. D. (1973). Enzymatic end-to-end joining of DNA molecules. J. Mol. Biol. 78: 453-471.
5. Casjens, S. (1979). Molecular organization of the bacteriophage P22 coat protein shell. J. Mol. Biol. 131:1-14.
6. Zhang, Z., Greene, B., Thuman-Commike, P., Jakana, J., Prevelige, P. Jr., King, J., and Chiu, W. (2000). Visualization of the maturation transition in bacteriophage P22 by electron cryomicroscopy. J. Mol. Biol. 297:615-26.
7. Steinbacher, S., Baxa, U., Miller, S., Weintraub, A., Seckler, R., and Huber, R. (1996). Crystal structure of phage P22 tailspike protein complexed with Salmonella sp. O-antigen receptors. Proc. Natl. Acad. Sci. USA 93:10584-8.
8. Casjens, S., and Hayden, M. (1988). Analysis in vivo of the bacteriophage P22 headful nuclease. J. Mol. Biol., 189:467-474.
9. Streisinger, G. Edgar, R., and Stahl, M. (1967). Chromosome structure in phage T4. Circularity of the linkage map. Proc. Natl. Acad. Sci., USA 57: 775-779.
10. Casjens, S., Wyckoff, E., Hayden, M., Sampson, L., Eppler, K., Randall, S., Moreno, E., and Serwer, P. (1992). The bacteriophage P22 portal protein is part of the gauge that determines the length and packing density of intravirion DNA. J. Mol. Biol. 224: 1055-1074.
11. Susskind, M., and Botstein, D. (1978) Molecular genetics of bacteriophage P22. Microbiol. Rev. 42: 385-413.
12. King J., Lenk E., Botstein D. (1973). Mechanism of head assembly and DNA encapsulation in Salmonella
13. King, J., and Casjens, S. (1974). Catalytic head assembling protein in virus morphogenesis. Nature 251:112-119.
14. S. Casjens and R. Hendrix, (1988) "Control mechanisms in dsDNA bacteriophage assembly", in The Bacteriophages, volume 1, ed. R. Calendar, Plenum Press, p. 15-91.
15. Wyckoff, E., and Casjens, S. (1985). Autoregulation of the bacteriophage P22 scaffolding protein gene. J. Virol. 53:192-197. phage P22. II. Morphogenetic pathway. J. Mol Biol. 80:697-731.
- A Brief Introduction to Bacteriophage P22 Assembly
Barrie Greene
- Visualization of the maturation transition in bacteriophage P22 by electron cryomicroscopy
J Mol Biol. 2000 Mar 31;297(3):615-26.
Zhang Z, Greene B, Thuman-Commike PA, Jakana J, Prevelige PE Jr, King J, Chiu W.
Verna and Marrs McLean Department of Biochemistry, Baylor College of Medicine, One Baylor Plaza, Houston, TX, 77030, USA.
Large-scale conformational transitions are involved in the life-cycle of many types of virus. The dsDNA phages, herpesviruses, and adenoviruses must undergo a maturation transition in the course of DNA packaging to convert a scaffolding-containing precursor capsid to the DNA-containing mature virion. This conformational transition converts the procapsid, which is smaller, rounder, and displays a distinctive skewing of the hexameric capsomeres, to the mature virion, which is larger and more angular, with regular hexons. We have used electron cryomicroscopy and image reconstruction to obtain 15 A structures of both bacteriophage P22 procapsids and mature phage. The maturation transition from the procapsid to the phage results in several changes in both the conformations of the individual coat protein subunits and the interactions between neighboring subunits. The most extensive conformational transformation among these is the outward movement of the trimer clusters present at all strict and local 3-fold axes on the procapsid inner surface. As the trimer tips are the sites of scaffolding binding, this helps to explain the role of scaffolding protein in regulating assembly and maturation. We also observe DNA within the capsid packed in a manner consistent with the spool model. These structures allow us to suggest how the binding interactions of scaffolding and DNA with the coat shell may act to control the packaging of the DNA into the expanding procapsids.
- Analysis in vivo of the bacteriophage P22 headful nuclease
J Mol Biol. 1988 Feb 5;199(3):467-74.
Casjens S, Hayden M.
Department of Cellular, Viral and Molecular Biology, University of Utah Medical Center, Salt Lake City 84132.
Bacteriophage P22 packages its double-stranded DNA chromosomes from concatemeric replicating DNA in a processive, sequential fashion. According to this model, during the initial packaging event in such a series the packaging apparatus recognizes a nucleotide sequence, called pac, on the DNA, and then condenses DNA within the coat protein shell unidirectionally (rightward) from that point. DNA ends are generated near the pac site before or during the condensation reaction. The right end of the mature chromosome is created by a cut made in the DNA by the "headful nuclease" after a complete chromosome is condensed within the phage head. Subsequent packaging events on that concatemeric DNA begin at the end generated by the headful cut of the previous event and proceed in the same direction as the previous event. We report here accurate measurements of the P22 chromosome length (43,400( +/- 750) base-pairs, where the uncertainty is the range in observed lengths), genome length (41,830( +/- 315) base-pairs, where the uncertainty represents the accuracy with which the length is known), the terminal redundancy (1600( +/- 750) base-pairs or 3.8( +/- 1.8)%, where the uncertainty is the observed range) and the imprecision in the headful measuring device ( +/- 750 base-pairs or +/- 1.7%). In addition, we present evidence for a weak nucleotide sequence specificity in the headful nuclease. These findings lend further support to, and extend our understanding of, the sequential series model of P22 DNA packaging.
- Visualization of the maturation transition in bacteriophage P22 by electron cryomicroscopy.
J Mol Biol. 2000 Mar 31;297(3):615-26.
Zhang Z, Greene B, Thuman-Commike PA, Jakana J, Prevelige PE Jr, King J, Chiu W.
Verna and Marrs McLean Department of Biochemistry, Baylor College of Medicine, One Baylor Plaza, Houston, TX, 77030, USA.
Large-scale conformational transitions are involved in the life-cycle of many types of virus. The dsDNA phages, herpesviruses, and adenoviruses must undergo a maturation transition in the course of DNA packaging to convert a scaffolding-containing precursor capsid to the DNA-containing mature virion. This conformational transition converts the procapsid, which is smaller, rounder, and displays a distinctive skewing of the hexameric capsomeres, to the mature virion, which is larger and more angular, with regular hexons. We have used electron cryomicroscopy and image reconstruction to obtain 15 A structures of both bacteriophage P22 procapsids and mature phage. The maturation transition from the procapsid to the phage results in several changes in both the conformations of the individual coat protein subunits and the interactions between neighboring subunits. The most extensive conformational transformation among these is the outward movement of the trimer clusters present at all strict and local 3-fold axes on the procapsid inner surface. As the trimer tips are the sites of scaffolding binding, this helps to explain the role of scaffolding protein in regulating assembly and maturation. We also observe DNA within the capsid packed in a manner consistent with the spool model. These structures allow us to suggest how the binding interactions of scaffolding and DNA with the coat shell may act to control the packaging of the DNA into the expanding procapsids.
- The structure of an infectious P22 virion shows the signal for headful DNA packaging.
Science. 2006 Jun 23;312(5781):1791-5. Epub 2006 May 18.
Lander GC, Tang L, Casjens SR, Gilcrease EB, Prevelige P, Poliakov A, Potter CS, Carragher B, Johnson JE.
Department of Molecular Biology, The Scripps Research Institute, 10550 North Torrey Pines Road, La Jolla, CA 92037, USA.
Bacteriophages, herpesviruses, and other large double-stranded DNA (dsDNA) viruses contain molecular machines that pump DNA into preassembled procapsids, generating internal capsid pressures exceeding, by 10-fold, that of bottled champagne. A 17 angstrom resolution asymmetric reconstruction of the infectious P22 virion reveals that tightly spooled DNA about the portal dodecamer forces a conformation that is significantly different from that observed in isolated portals assembled from ectopically expressed protein. We propose that the tight dsDNA spooling activates the switch that signals the headful chromosome packing density to the particle exterior.
J Mol Biol. 2007 Aug 10;371(2):374-87. Epub 2007 May 24.
Department of Biochemistry and Molecular Biology, SUNY Upstate Medical University, 750, E. Adams Street, Syracuse, NY 13210, USA.
The attachment of tailed bacteriophages to the host cell wall as well as the penetration and injection of the viral genome into the host is mediated by the virion tail complex. In phage P22, a member of the Podoviridae family that infects Salmonella enterica, the tail contains an approximately 220 A elongated protein needle, previously identified as tail accessory factor gp26. Together with tail factors gp4 and gp10, gp26 is critical to close the portal protein channel and retain the viral DNA inside the capsid. By virtue of its topology and position in the virion, the tail needle gp26 is thought to function as a penetrating device to perforate the Salmonella cell wall. Here, we define the domain organization of gp26, characterize the structural determinants for its stability, and define the polarity of the gp26 assembly into the phage portal vertex structure. We have found that the N-terminal 27 residues of gp26 form a functional domain that, although not required for gp26 trimerization and overall stability, is essential for the correct attachment to gp10, which is thought to plug the portal vertex structure. The region downstream of domain I, domain II, folds into helical core, which exhibits four trimerization octad repeats with consensus Ile-xx-Leu-xxx-Val/Tyr. We demonstrate that in vitro, domain II represents the main self-assembling, highly stable trimerization core of gp26, which retains a folded conformation both in an anhydrous environment and in the presence of 10% SDS. The C terminus of gp26, immediately downstream of domain II, contains a beta-sheet-rich region, domain III, and a short coiled coil, domain IV, which, although not required for gp26 trimerization, enhance its thermodynamic stability. We propose that domains III and IV of the tail needle form the tip utilized by the phage to penetrate the host cell wall.
P22 tail machine
Upon encountering a host bacterium, the tail section of the virion binds to receptors on the cell surface, and delivers the DNA into the cell by use of an injectisome like mechanism (an injectisome is a nanomachine that evolved for the delivery of proteins, by type III secretion). The tail section of the virus punches a hole through the bacterial cell wall and plasma membrane, and the genome passes down the tail into the cell.
from http://carbon.bio.ku.edu/gallery.htm:
Bacteriophage P22
A 3D model of P22 virion derived from cryoEM maps of the phage head and tail.
P22 tail machine
CryoEM map of a 5-protein, 51-subunit molecular machine
P22 portal
8 A resolution cryoEM map fitted with X-ray structure of phi29 portal
- Three-dimensional structure of the bacteriophage P22 tail machine
Liang Tang, William R Marion, Gino Cingolani, Peter E Prevelige Jr and John E Johnson
The EMBO Journal (2005) 24, 2087–2095
The tail of the bacteriophage P22 is composed of multiple protein components and integrates various biological functions that are crucial to the assembly and infection of the phage. The three-dimensional structure of the P22 tail machine determined by electron cryo-microscopy and image reconstruction reveals how the five types of polypeptides present as 51 subunits are organized into this molecular machine through twelve-, six- and three-fold symmetry, and provides insights into molecular events during host cell attachment and phage DNA translocation.
- Structure and Functions of the Bacteriophage P22 Tail Protein
Peter B. Berget and Anthony R. Poteete (MIT),
Journal of Virology, 1980 April; 34(1): 234-243
The product of gene 9 (gp9) of Salmonella typhimurium bacteriophage P22 is a multifunctional structural protein. This protein is both a specific glycosidase which imparts the adsorption characteristics of the phage for its host and a protein which participates in a specific assembly reaction during phage morphogenesis. We have begun a detailed biochemical and genetic analysis of this gene product. A relatively straightforward purification of this protein has been devised, and various physical parameters of the protein have been determined. The protein has an s20,w of 9.3S, a D20,w of 4.3 x 10–7 cm2/s, and a molecular weight, as determined by sedimentation equilibrium, of 173,000. The purified protein appears as a prolate ellipsoid upon electron microscopic examination, with an axial ratio of 4:1, which is similar to the observed shape when it is attached to the phage particle. The molecular weight is consistent with the tail protein being a dimer of gp9 and each phage containing six of these dimers. An altered form of the tail protein has been purified from supF cells infected with a phage strain carrying an amber mutation in gene 9. Phage "tailed" with this altered form of gp9 adsorb to susceptible cells but form infectious centers with a severely reduced efficiency (ca. 1%). Biochemical analysis of the purified wild-type and genetically altered tail proteins suggests that loss of infectivity correlates with a loss in the glycosidase activity of the protein (2.5% residual activity). From these results we propose that the glycosidic activity of the P22 tail protein is not essential for phage assembly or adsorption of the phage to its host but is required for subsequent steps in the process of infection.
P22 capsid
- Molecular assembly of bacteriophage P22
Also see "Penton release from P22 heat-expanded capsids suggests importance of stabilizing penton-hexon interactions during capsid maturation." Biophysical Journal 84, 2585-2592, Teschke, C. M., McGough, A. & Thuman-Commike, P. A. (2003).
[ These describe the capsid without hte 5-fold end-cap. This would be interesting to compare directly with my segmented P22 vertex. ]
Assembly of an icosahedral virus must proceed with exquisite fidelity, and is a paradigm for the self-organization of multiple subunits required to form complex macromolecular structures. We investigate virus assembly using the bacteriophage P22. In phage P22, herpesvirus and many other dsDNA viruses, the initial product of the assembly reaction is a precursor capsid, known as the procapsid 1. Scaffolding protein directs proper assembly of coat protein, the major capsid protein, to form the procapsid 1. The procapsid undergoes expansion during DNA packaging to become the mature virion 2; 3; 4. P22 assembly is an excellent model system because these complex in vivo processes can be mimicked in vitro.
P22 procapsids can be induced to undergo the structural transformation that accompanies capsid maturation by heat or chemical treatment. We examined bacteriophage P22 morphogenesis by comparing three-dimensional structures of capsids expanded both in vitro by heat treatment and in vivo by DNA packaging 8. The structures were determined by electron cryo-microscopy to 24 Å.
Figure 2. Outer surface representation of (A) the heat expanded capsid and (B) the in vivo expanded empty head three-dimensional reconstructions viewed from the icosahedral three-fold axis. The unit triangle and one hexon have been outlined on each capsid for a point of reference.
Assymmetric reconstruction of Epsilon 15
Biologists build better software, beat path to viral knowledge
Wen Jiang, Juan Chang, Joanita Jakana, Peter Weigele, Jonathan King and Wah Chiu
Nature 439, 612-616 (2 February 2006) | doi:10.1038/nature04487
The critical viral components for packaging DNA, recognizing and binding to host cells, and injecting the condensed DNA into the host are organized at a single vertex of many icosahedral viruses. These component structures do not share icosahedral symmetry and cannot be resolved using a conventional icosahedral averaging method. Here we report the structure of the entire infectious Salmonella bacteriophage epsilon15 (ref. 1) determined from single-particle cryo-electron microscopy, without icosahedral averaging. This structure displays not only the icosahedral shell of 60 hexamers and 11 pentamers, but also the non-icosahedral components at one pentameric vertex. The densities at this vertex can be identified as the 12-subunit portal complex sandwiched between an internal cylindrical core and an external tail hub connecting to six projecting trimeric tailspikes. The viral genome is packed as coaxial coils in at least three outer layers with90 terminal nucleotides extending through the protein core and the portal complex and poised for injection. The shell protein from icosahedral reconstruction at higher resolution exhibits a similar fold to that of other double-stranded DNA viruses including herpesvirus, suggesting a common ancestor among these diverse viruses. The image reconstruction approach should be applicable to studying other biological nanomachines with components of mixed symmetries.
Physics and structure of DNA packaging
- The Bacteriophage DNA Packaging Motor
Annual Review of Genetics
Vol. 42 (Volume publication date December 2008)
Venigalla B. Rao Department of Biology, The Catholic University of America,Washington, D.C. 20064; email: rao@cua.eduMichael Feiss Department of Microbiology, Roy J. and Lucille A. Carver College of Medicine, University of Iowa, Iowa City, Iowa 52242; email: michael-feiss@uiowa.edu An ATP-powered DNA translocation machine encapsidates the viral genome in the large dsDNA bacteriophages. The essential components include the empty shell, prohead, and the packaging enzyme, terminase. During translocation, terminase is docked on the prohead's portal protein. The translocation ATPase and the concatemer-cutting endonuclease reside in terminase. Remarkably, terminases, portal proteins, and shells of tailed bacteriophages and herpesviruses show conserved features. These DNA viruses may have descended from a common ancestor. Terminase's ATPase consists of a classic nucleotide binding fold, most closely resembling that of monomeric helicases. Intriguing models have been proposed for the mechanism of dsDNA translocation, invoking ATP hydrolysis-driven conformational changes of portal or terminase powering DNA motion. Single-molecule studies show that the packaging motor is fast and powerful. Recent advances permit experiments that can critically test the packaging models. The viral genome translocation mechanism is of general interest, given the parallels between terminases, helicases, and other motor proteins.
- DNA packaging and ejection forces in bacteriophage
James Kindt†, Shelly Tzlil‡, Avinoam Ben-Shaul‡, and William M. Gelbart†
†Department of Chemistry and Biochemistry, University of California, Los Angeles, CA 90095-1569; and ‡Department of Physical Chemistry and The Fritz Haber Research Center, Hebrew University, Jerusalem 91904, Israel
We calculate the forces required to package (or, equivalently, acting to eject) DNA into (from) a bacteriophage capsid, as a function of the loaded (ejected) length, under conditions for which the DNA is either self-repelling or self-attracting. Through computer simulation and analytical theory, we find the loading force to increase more than 10-fold (to tens of piconewtons) during the final third of the loading process; correspondingly, the internal pressure drops 10-fold to a few atmospheres (matching the osmotic pressure in the cell) upon ejection of just a small fraction of the phage genome. We also determine an evolution of the arrangement of packaged DNA from toroidal to spool-like structures.
- Dynamics of DNA Ejection from Bacteriophage
Biophysical Journal 91:411-420 (2006)
Mandar M. Inamdar * ,William M. Gelbart andRob Phillips * * Division of Engineering and Applied Science,Kavli Nanoscience Institute, California Institute of Technology, Pasadena, California; and
Department of Chemistry and Biochemistry, University of California, Los Angeles, California
The ejection of DNA from a bacterial virus (i.e., phage) into its host cell is a biologically important example of the translocation of a macromolecular chain along its length through a membrane. The simplest mechanism for this motion is diffusion, but in the case of phage ejection a significant driving force derives from the high degree of stress to which the DNA is subjected in the viral capsid. The translocation is further sped up by the ratcheting and entropic forces associated with proteins that bind to the viral DNA in the host cell cytoplasm. We formulate a generalized diffusion equation that includes these various pushing and pulling effects and make estimates of the corresponding speedups in the overall translocation process. Stress in the capsid is the dominant factor throughout early ejection, with the pull due to binding particles taking over at later stages. Confinement effects are also investigated, in the case where the phage injects its DNA into a volume comparable to the capsid size. Our results suggest a series of in vitro experiments involving the ejection of DNA into vesicles filled with varying amounts of binding proteins from phage whose state of stress is controlled by ambient salt conditions or by tuning genome length.
- Osmotic Pressure and Packaging Structure of Caged DNA
Biophysical Journal 94:737-746 (2008)
Zhidong Li * ,Jianzhong Wu * andZhen-Gang Wang * Department of Chemical and Environmental Engineering, University of California, Riverside, California,
We present a theoretical model for aqueous solutions of double-stranded (ds) DNA with explicit consideration of electrostatic interactions, excluded-volume effects, van der Waals attractions, and salt ions. With reasonable parameters estimated from the DNA structure and experimental data for electrolytes, we are able to reproduce the DNA osmotic pressure in the bulk in good agreement with experiment. The predicted DNA osmotic pressure in
-bacteriophages is found to coincide with that of the PEG8000 solution that inhibits DNA ejection as reported in recent experiments. Based on the radial distributions of DNA segments and of counterions at different degrees of packaging, we find that in the presence of Mg2+, DNA forms a multilayer structure near the inner surface of a fully loaded bacteriophage, but at low packing density the DNA segments are depleted from the surface owing to the local condensation of DNA induced by the divalent counterions. By contrast, the multilayer DNA structure is less distinctive in the presence of Na+ despite the increase of the DNA density at contact, and the depletion near the capsid surface is not found at low packing density.
DNA packaging molecular motors
- Clockwork That Drives Powerful Virus Nanomotor Discovered , from ScienceDaily , Dec. 31, 2008
"Not all viruses have a motor such as the one found in the T4 virus, but some viruses that cause human diseases posses molecular motors with similar functions, and likely have similar structures. T4 uses its motor to pack about 171,000 basepairs of genetic information to near-crystalline density within its 120 nanometer by 86 nanometer capsid.
The researchers found that the motor is located at the intersection of the capsid and the virus "tail" and is made of a circular array of proteins called gene product 17 (gp17). Five, two-part, gp17 proteins combine to form a pair of conjoined rings, arrayed so that their upper segments form an upper ring and their lower segments form a lower ring.
As a T4 virus assembles itself, the lower ring of the motor structure attaches to a strand of DNA, while the upper ring attaches to a capsid. The upper and lower rings have opposite charges, which allow the motor to contract and release, alternately tugging at the DNA like a ring of hands pulling on a rope."
- From Wikipedia Molecular motor:
Molecular motors are biological molecular machines that are the essential agents of movement in living organisms. Generally speaking, a motor may be defined as a device that consumes energy in one form and converts it into motion or mechanical work; for example, many protein-based molecular motors harness the chemical free energy released by the hydrolysis of ATP in order to perform mechanical work [1]. In terms of energetic efficiency, these types of motors can be superior to currently available man-made motors. One important difference between molecular motors and macroscopic motors is that molecular motors operate in the thermal bath, an environment where the fluctuations due to thermal noise are significant.
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Recently, chemists and those involved in nanotechnology have begun to explore the possibility of creating molecular motors de novo. These synthetic molecular motors currently suffer many limitations that confine their use to the research laboratory. However, many of these limitations may be overcome as our understanding of chemistry and physics at the nanoscale increases. Systems like the nanocars, while not technically motors, are illustrative of recent efforts towards synthetic nanoscale motors.
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Viral DNA packaging motors inject viral genomic DNA into capsids as part of their replication cycle, packing it very tightly. [5]
- From Wikipedia Motor protien:
Motor proteins are a class of molecular motors that are able to move along the surface of a suitable substrate. They are powered by the hydrolysis of ATP and convert chemical energy into mechanical work.
The most prominent example of a motor protein is the muscle protein myosin which "motors" the contraction of muscle fibers in animals. Motor proteins are the driving force behind most active transport of proteins and vesicles in the cytoplasm. Kinesins and dyneins play essential roles in intracellular transport such as axonal transport and in the formation of the spindle apparatus and the separation of the chromosomes during mitosis and meiosis. Dynein is found in flagella and is crucial to cell motility, for example in spermatozoa
Derek N. Fuller*, Dorian M. Raymer*, Vishal I. Kottadiel†, Venigalla B. Rao†‡, and Douglas E. Smith*‡
*Department of Physics, University of California at San Diego, La Jolla, CA 92093; and †Department of Biology, The Catholic University of America,
Washington, DC 20064Terminase enzyme complexes, which facilitate ATP-driven DNA
packaging in phages and in many eukaryotic viruses, constitute a
wide and potentially diverse family of molecular motors about
which little dynamic or mechanistic information is available. Here
we report optical tweezers measurements of single DNA molecule
packaging dynamics in phage T4, a large, tailed Escherichia coli
virus that is an important model system in molecular biology. We
show that a complex is formed between the empty prohead and
the large terminase protein (gp17) that can capture and begin
packaging a target DNA molecule within a few seconds, thus
demonstrating a distinct viral assembly pathway. The motor generates
forces >60 pN, similar to those measured with phage 29,
suggesting that high force generation is a common property of
viral DNA packaging motors. However, the DNA translocation rate
for T4 was strikingly higher than that for29, averaging700 bp/s
and ranging up to 2,000 bp/s, consistent with packaging by
phage T4 of an enormous, 171-kb genome in <10 min during viral
infection and implying high ATP turnover rates of >300 s1. The
motor velocity decreased with applied load but averaged 320 bp/s
at 45 pN, indicating very high power generation. Interestingly, the
motor also exhibited large dynamic changes in velocity, suggesting
that it can assume multiple active conformational states gearing
different translocation rates. This capability, in addition to the
reversible pausing and slipping capabilities that were observed,
may allow phage T4 to coordinate DNA packaging with other
ongoing processes, including viral DNA transcription, recombination,
and repair.
Structure of the Bacteriophage phi29 DNA Packaging Motor
Nature, December 7, 2000
Alan A. Simpson, Yizhi Tao, Petr G. Leiman, Mohammed O. Badasso, Yongning He, Paul J. Jardine, Norman H. Olson, Marc C. Morais, Shelley Grimes, Dwight L. Anderson, Timothy S. Baker, and Michael G. Rossmann
Motors generating mechanical force, powered by the hydrolysis of ATP, are used to translocate double-stranded DNA (dsDNA) into preformed capsids (proheads) of bacterial viruses and certain animal viruses. Here, we describe the motor that packages the dsDNA of the Bacillus subtilis bacteriophage phi29 into a precursor capsid. The structure of the head-tail connector, the central component of the phi29 DNA packaging motor, was determined to 3.2 angstrom resolution by means of X-ray crystallography. The connector was fitted into the electron density of the prohead and the partially packaged prohead determined by cryo-electron microscopy (cryoEM) image reconstructions. Our results suggest that the prohead plus dodecameric connector, prohead RNA (pRNA), viral ATPase, and DNA comprise a rotary motor with the head-pRNA-ATPase complex acting as a stator, the DNA acting as a spindle, and the connector as a ball-race. The helical nature of the DNA converts the rotary action of the connector into translation of the DNA.
Link to Quicktime Movie of DNA packaging motorDerek N. Fuller, Dorian M. Raymer, Vishal I. Kottadiel, Venigalla B. Rao, and Douglas E. Smith
PNAS October 23, 2007 vol. 104 no. 43 16868-16873
Terminase enzyme complexes, which facilitate ATP-driven DNA packaging in phages and in many eukaryotic viruses, constitute a wide and potentially diverse family of molecular motors about which little dynamic or mechanistic information is available. Here we report optical tweezers measurements of single DNA molecule packaging dynamics in phage T4, a large, tailed Escherichia coli virus that is an important model system in molecular biology. We show that a complex is formed between the empty prohead and the large terminase protein (gp17) that can capture and begin packaging a target DNA molecule within a few seconds, thus demonstrating a distinct viral assembly pathway. The motor generates forces >60 pN, similar to those measured with phage φ29, suggesting that high force generation is a common property of viral DNA packaging motors. However, the DNA translocation rate for T4 was strikingly higher than that for φ29, averaging ≈700 bp/s and ranging up to ≈2,000 bp/s, consistent with packaging by phage T4 of an enormous, 171-kb genome in <10 min during viral infection and implying high ATP turnover rates of >300 s−1. The motor velocity decreased with applied load but averaged 320 bp/s at 45 pN, indicating very high power generation. Interestingly, the motor also exhibited large dynamic changes in velocity, suggesting that it can assume multiple active conformational states gearing different translocation rates. This capability, in addition to the reversible pausing and slipping capabilities that were observed, may allow phage T4 to coordinate DNA packaging with other ongoing processes, including viral DNA transcription, recombination, and repair.
- DNA packaging and delivery machines in tailed bacteriophages.
Curr Opin Struct Biol. 2007 Apr;17(2):237-43. Epub 2007 Mar 28.
Department of Molecular Biology, The Scripps Research Institute, La Jolla, CA 92037, USA. jackj@scripps.edu
Several symmetric and asymmetric reconstructions of bacteriophage particles have recently been determined using electron cryo-microscopy and image reconstruction, and X-ray crystal structures of phage particles and particle-associated gene products have also been solved. In the past two years, the asymmetric structures of four different phages, T7, epsilon15, P22 and phi29, were determined at resolutions sufficient to visualize details of the machinery for DNA packaging and delivery, as well as the organization of the double-stranded DNA within the particles. Invariably, the portals, through which DNA enters and leaves the particle, have 12-fold symmetry, occupy a pentavalent site in the capsid and, along with tail machine accessory proteins attached to it, are fixed in a specific orientation relative to the rest of the capsid.
- DNA Packaging by Bacteriophage P22
Chapter 5 of "Viral Genome Packaging Machines: Genetics, Structure, and Mechanism", Sherwood Casjens and Peter Weigele, 2005
- Plant Virus Research Could Lead To New Ways To Improve Crop Yields
A 3D representation of the structure of the protein coat for the soybean mosaic virus showing an exterior view, axial and longitudinal cross section. (Credit: Image courtesy of Stubbs Lab, Vanderbilt University)
ScienceDaily (Oct. 9, 2008) — An interdisciplinary group of scientists has obtained the first detailed information about the structure of the most destructive group of plant viruses known: flexible filamentous viruses.
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The viruses are much too small to see with an optical microscope. So the scientists had to use a combination of imaging techniques — cryo-electron microscopy at Vanderbilt, X-ray diffraction at Argonne and scanning transmission electron microscopy (STEM) at Brookhaven – in order to identify the two structures.
“These techniques are very complementary,” says Stubbs. “People have been trying to get this structural work started for decades, more than 40 years. It’s been very difficult and there have been a number of obstacles, including the fact that it’s very hard to make good samples of these viruses.”
Even after Stubbs and his colleagues figured out how to make good samples and analyzed them using X-ray diffraction and traditional electron microscopy, there were a number of ambiguities in the results. Brookhaven’s STEM technique provided the definitive answers.
Although it could not determine the structure of the viral protein coat directly, STEM was able to put boundaries on the number of molecules in each "turn" of the spiral-shaped structure. This allowed the scientists to pick the correct structure from the alternatives they had identified using the other techniques.
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