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The DNA-packaging nanomotor of tailed bacteriophages

Key Points

  • Tailed bacteriophages package their double-stranded DNA chromosomes into preformed protein containers called procapsids.

  • The DNA-packaging process condenses the DNA several-hundred-fold and requires chemical energy. The result of this condensation is DNA that is arranged as a tightly wound, imperfect solenoid that is held within the virion with no proteins holding the DNA strands close together.

  • The molecular motor that packages DNA within the procapsid is a DNA translocase that is composed of three proteins: portal protein, which forms the hole through which DNA enters the procapsid, and also senses when the capsid is full of DNA; small terminase subunit (TerS), which recognizes the DNA that will be packaged; and large terminase subunit (TerL), the ATPase that converts the chemical energy of ATP hydrolysis into physical motion of the DNA.

  • In the phages that package over-length or circular DNA, TerL also contains a nuclease activity that cuts the DNA to form the unit-length, linear DNA molecules that are found in completed virions.

  • Single-molecule experiments have shown that the DNA-packaging motor can insert DNA into the procapsid at rates of up to 1,800 bp per second, generates forces of up to 100 piconewtons, and translocates the DNA in steps of 10 bp that in turn consist of four rapid 2.5-bp substeps.

  • TerL and probably TerS are physical components of the packaging motor that are released from the nascent virion upon completion of packaging, so they are not present in completed virions.

  • After DNA packaging and TerL release, other proteins stabilize the packaged DNA by plugging the channel through which the DNA entered the procapsid and by binding the outside of the capsid shell to strengthen it.

  • Recently obtained atomic structures of packaging motor proteins, channel plug proteins and shell-strengthening proteins, as well as subnanometer-resolution, cryo-electron microscopy-derived three-dimensional reconstructions of virions and procapsids, have shed new light on the mechanism of the motor and its control.

Abstract

Tailed bacteriophages use nanomotors, or molecular machines that convert chemical energy into physical movement of molecules, to insert their double-stranded DNA genomes into virus particles. These viral nanomotors are powered by ATP hydrolysis and pump the DNA into a preformed protein container called a procapsid. As a result, the virions contain very highly compacted chromosomes. Here, I review recent progress in obtaining structural information for virions, procapsids and the individual motor protein components, and discuss single-molecule in vitro packaging reactions, which have yielded important new information about the mechanism by which these powerful molecular machines translocate DNA.

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Figure 1: Tailed bacteriophages and their DNA-packaging motor.
Figure 2: Arrangement of DNA and the portal protein in bacteriophage P22 virions.
Figure 3: Structures of the three DNA-packaging motor proteins.
Figure 4: The procapsid of phage P22.
Figure 5: DNA-packaging motor of phage T4.

References

  1. Bergh, O., Borsheim, K. Y., Bratbak, G. & Heldal, M. High abundance of viruses found in aquatic environments. Nature 340, 467–468 (1989).

    CAS  PubMed  Google Scholar 

  2. Suttle, C. A. Viruses in the sea. Nature 437, 356–361 (2005).

    CAS  PubMed  Article  Google Scholar 

  3. Whitman, W. B., Coleman, D. C. & Wiebe, W. J. Prokaryotes: the unseen majority. Proc. Natl Acad. Sci. USA 95, 6578–6583 (1998).

    CAS  Article  PubMed  PubMed Central  Google Scholar 

  4. Wommack, K. E. & Colwell, R. R. Virioplankton: viruses in aquatic ecosystems. Microbiol. Mol. Biol. Rev. 64, 69–114 (2000).

    CAS  PubMed  PubMed Central  Article  Google Scholar 

  5. Brussow, H. & Hendrix, R. W. Phage genomics: small is beautiful. Cell 108, 13–16 (2002).

    CAS  PubMed  Article  Google Scholar 

  6. Boyd, E. F., Davis, B. M. & Hochhut, B. Bacteriophage–bacteriophage interactions in the evolution of pathogenic bacteria. Trends Microbiol. 9, 137–144 (2001).

    CAS  PubMed  Article  Google Scholar 

  7. Casjens, S. & Hendrix, R. in The Bacterial Chromosome (ed. Higgins, N. P.) 39–52 (American Society for Microbiology Press, Washingtion DC, 2005).

    Book  Google Scholar 

  8. Cheetham, B. F. & Katz, M. E. A role for bacteriophages in the evolution and transfer of bacterial virulence determinants. Mol. Microbiol. 18, 201–208 (1995).

    CAS  PubMed  Article  Google Scholar 

  9. Denou, E. et al. T4 phages against Escherichia coli diarrhea: potential and problems. Virology 388, 21–30 (2009).

    CAS  PubMed  Article  Google Scholar 

  10. Summers, W. C. Bacteriophage therapy. Annu. Rev. Microbiol. 55, 437–451 (2001).

    CAS  PubMed  Article  Google Scholar 

  11. Luftig, R. B., Wood, W. B. & Okinaka, R. Bacteriophage T4 head morphogenesis. On the nature of gene 49-defective heads and their role as intermediates. J. Mol. Biol. 57, 555–573 (1971).

    CAS  PubMed  Article  Google Scholar 

  12. Kaiser, D., Syvanen, M. & Masuda, T. DNA packaging steps in bacteriophage lambda head assembly. J. Mol. Biol. 91, 175–186 (1975).

    CAS  PubMed  Article  Google Scholar 

  13. Catalano, C. (ed.) Viral Genome Packaging Machines: Genetics, Structure, and Mechanism (Landes Bioscience, Georgetown, Texas, 2005).

    Book  Google Scholar 

  14. Casjens, S. & Hendrix, R. in The Bacteriophages (ed. Calendar, R.) 15–91 (Plenum, New York, 1988).

    Book  Google Scholar 

  15. Earnshaw, W. & Casjens, S. DNA packaging by the double-stranded DNA bacteriophages. Cell 21, 319–331 (1980).

    CAS  PubMed  Article  Google Scholar 

  16. Black, L. W. DNA packaging in dsDNA bacteriophages. Annu. Rev. Microbiol. 43, 267–292 (1989).

    CAS  PubMed  Article  Google Scholar 

  17. Maluf, N. K. & Feiss, M. Virus DNA translocation: progress towards a first ascent of Mount Pretty Difficult. Mol. Microbiol. 61, 1–4 (2006).

    CAS  PubMed  Article  Google Scholar 

  18. Rao, V. B. & Feiss, M. The bacteriophage DNA packaging motor. Annu. Rev. Genet. 42, 647–681 (2008).

    CAS  PubMed  Article  Google Scholar 

  19. Jeembaeva, M., Jonsson, B., Castelnovo, M. & Evilevitch, A. DNA heats up: energetics of genome ejection from phage revealed by isothermal titration calorimetry. J. Mol. Biol. 395, 1079–1087 (2010).

    CAS  PubMed  Article  Google Scholar 

  20. Panja, D. & Molineux, I. J. Dynamics of bacteriophage genome ejection in vitro and in vivo. Phys. Biol. 7, 045006 (2010).

    PubMed  Article  Google Scholar 

  21. Kindt, J., Tzlil, S., Ben-Shaul, A. & Gelbart, W. M. DNA packaging and ejection forces in bacteriophage. Proc. Natl Acad. Sci. USA 98, 13671–13674 (2001).

    CAS  PubMed  Article  PubMed Central  Google Scholar 

  22. Riemer, S. C. & Bloomfield, V. A. Packaging of DNA in bacteriophage heads: some considerations on energetics. Biopolymers 17, 785–794 (1978).

    CAS  PubMed  Article  Google Scholar 

  23. Agirrezabala, X. et al. Maturation of phage T7 involves structural modification of both shell and inner core components. EMBO J. 24, 3820–3829 (2005).

    CAS  PubMed  PubMed Central  Article  Google Scholar 

  24. Choi, K. H. et al. Insight into DNA and protein transport in double-stranded DNA viruses: the structure of bacteriophage N4. J. Mol. Biol. 378, 726–736 (2008).

    CAS  PubMed  PubMed Central  Article  Google Scholar 

  25. Jiang, W. et al. Structure of epsilon15 bacteriophage reveals genome organization and DNA packaging/injection apparatus. Nature 439, 612–616 (2006).

    CAS  PubMed  PubMed Central  Article  Google Scholar 

  26. Lander, G. C. et al. The structure of an infectious P22 virion shows the signal for headful DNA packaging. Science 312, 1791–1795 (2006).

    CAS  PubMed  Article  Google Scholar 

  27. Lander, G. C. et al. Bacteriophage lambda stabilization by auxiliary protein gpD: timing, location, and mechanism of attachment determined by cryo-EM. Structure 16, 1399–1406 (2008). This report describes the subnanometre, cryo-electron microscopy-derived three-dimensional reconstruction of phage λ capsids, with a focus on the bacteriophage shell-strengthening protein.

    CAS  PubMed  PubMed Central  Article  Google Scholar 

  28. Leiman, P. G. et al. The structures of bacteriophages K1E and K1-5 explain processive degradation of polysaccharide capsules and evolution of new host specificities. J. Mol. Biol. 371, 836–849 (2007).

    CAS  Article  PubMed  Google Scholar 

  29. Tang, J. et al. DNA poised for release in bacteriophage φ29. Structure 16, 935–943 (2008).

    CAS  PubMed  PubMed Central  Article  Google Scholar 

  30. Chang, J., Weigele, P., King, J., Chiu, W. & Jiang, W. Cryo-EM asymmetric reconstruction of bacteriophage P22 reveals organization of its DNA packaging and infecting machinery. Structure 14, 1073–1082 (2006).

    CAS  PubMed  Article  Google Scholar 

  31. Duda, R. L., Hendrix, R. W., Huang, W. M. & Conway, J. F. Shared architecture of bacteriophage SPO1 and herpesvirus capsids. Curr. Biol. 16, R11–R13 (2006).

    CAS  PubMed  Article  Google Scholar 

  32. Liu, X. et al. Structural changes in a marine podovirus associated with release of its genome into Prochlorococcus. Nature Struct. Mol. Biol. 17, 830–836 (2010).

    CAS  Article  Google Scholar 

  33. Fokine, A. et al. Molecular architecture of the prolate head of bacteriophage T4. Proc. Natl Acad. Sci. USA 101, 6003–6008 (2004).

    CAS  PubMed  Article  PubMed Central  Google Scholar 

  34. Effantin, G., Boulanger, P., Neumann, E., Letellier, L. & Conway, J. F. Bacteriophage T5 structure reveals similarities with HK97 and T4 suggesting evolutionary relationships. J. Mol. Biol. 361, 993–1002 (2006).

    CAS  PubMed  Article  Google Scholar 

  35. Dai, W. et al. Three-dimensional structure of tropism-switching Bordetella bacteriophage. Proc. Natl Acad. Sci. USA 107, 4347–4352 (2010).

    CAS  PubMed  Article  PubMed Central  Google Scholar 

  36. Cerritelli, M. E. et al. Encapsidated conformation of bacteriophage T7 DNA. Cell 91, 271–280 (1997).

    CAS  PubMed  Article  Google Scholar 

  37. Leforestier, A. & Livolant, F. Structure of toroidal DNA collapsed inside the phage capsid. Proc. Natl Acad. Sci. USA 106, 9157–9162 (2009).

    CAS  PubMed  Article  PubMed Central  Google Scholar 

  38. Agirrezabala, X. et al. Structure of the connector of bacteriophage T7 at 8A resolution: structural homologies of a basic component of a DNA translocating machinery. J. Mol. Biol. 347, 895–902 (2005).

    CAS  PubMed  Article  Google Scholar 

  39. Cerritelli, M. E. et al. A second symmetry mismatch at the portal vertex of bacteriophage T7, 8-fold symmetry in the procapsid core. J. Mol. Biol. 327, 1–6 (2003).

    CAS  PubMed  Article  Google Scholar 

  40. Tang, J. et al. Peering down the barrel of a bacteriophage portal: the genome packaging and release valve in P22. Structure 19, 496–502 (2011).

    CAS  PubMed  PubMed Central  Article  Google Scholar 

  41. Black, L. W. & Silverman, D. J. Model for DNA packaging into bacteriophage T4 heads. J. Virol. 28, 643–655 (1978).

    CAS  PubMed  PubMed Central  Google Scholar 

  42. de Frutos, M., Letellier, L. & Raspaud, E. DNA ejection from bacteriophage T5: analysis of the kinetics and energetics. Biophys. J. 88, 1364–1370 (2005).

    CAS  PubMed  Article  Google Scholar 

  43. Leforestier, A. & Livolant, F. The bacteriophage genome undergoes a succession of intracapsid phase transitions upon DNA ejection. J. Mol. Biol. 396, 384–395 (2010).

    CAS  PubMed  Article  Google Scholar 

  44. Sao-Jose, C., de Frutos, M., Raspaud, E., Santos, M. A. & Tavares, P. Pressure built by DNA packing inside virions: enough to drive DNA ejection in vitro, largely insufficient for delivery into the bacterial cytoplasm. J. Mol. Biol. 374, 346–355 (2007).

    CAS  PubMed  Article  Google Scholar 

  45. Letellier, L., Boulanger, P., Plancon, L., Jacquot, P. & Santamaria, M. Main features on tailed phage, host recognition and DNA uptake. Front. Biosci. 9, 1228–1339 (2004).

    CAS  PubMed  Article  Google Scholar 

  46. Casjens, S. & Molineux, I. in Viral Molecular Machines (eds Rossmann, M. & Rao, V.) (Springer, New York, in the press).

  47. Casjens, S. R. Comparative genomics and evolution of the tailed-bacteriophages. Curr. Opin. Microbiol. 8, 451–458 (2005).

    CAS  PubMed  Article  Google Scholar 

  48. Hendrix, R. W. Bacteriophages: evolution of the majority. Theor. Popul. Biol. 61, 471–480 (2002).

    PubMed  Article  Google Scholar 

  49. Kochan, J., Carrascosa, J. L. & Murialdo, H. Bacteriophage lambda preconnectors: purification and structure. J. Mol. Biol. 174, 433–447 (1984).

    CAS  PubMed  Article  Google Scholar 

  50. Driedonks, R. A., Engel, A., tenHeggeler, B. & van Driel, R. Gene 20 product of bacteriophage T4 its purification and structure. J. Mol. Biol. 152, 641–662 (1981).

    CAS  PubMed  Article  Google Scholar 

  51. Carazo, J. M., Donate, L. E., Herranz, L., Secilla, J. P. & Carrascosa, J. L. Three-dimensional reconstruction of the connector of bacteriophage φ29 at 1.8 nm resolution. J. Mol. Biol. 192, 853–867 (1986).

    CAS  PubMed  Article  Google Scholar 

  52. Lurz, R. et al. Structural organisation of the head-to-tail interface of a bacterial virus. J. Mol. Biol. 310, 1027–1037 (2001).

    CAS  PubMed  Article  Google Scholar 

  53. Doan, D. N. & Dokland, T. The gpQ portal protein of bacteriophage P2 forms dodecameric connectors in crystals. J. Struct. Biol. 157, 432–436 (2007).

    CAS  PubMed  Article  Google Scholar 

  54. Morais, M. C. et al. Defining molecular and domain boundaries in the bacteriophage φ29 DNA packaging motor. Structure 16, 1267–1274 (2008). This investigation uses cryo-electron microscopy to obtain a three-dimensional reconstruction of the phage φ29 procapsid with the TerL ATPase bound to it.

    CAS  PubMed  PubMed Central  Article  Google Scholar 

  55. Simpson, A. A. et al. Structure of the bacteriophage φ29 DNA packaging motor. Nature 408, 745–750 (2000).

    CAS  PubMed  PubMed Central  Article  Google Scholar 

  56. Lebedev, A. A. et al. Structural framework for DNA translocation via the viral portal protein. EMBO J. 26, 1984–1994 (2007).

    CAS  PubMed  PubMed Central  Article  Google Scholar 

  57. Olia, A., Prevelige, P. E. Jr, Johnson, J. & Cingolani, G. Three-dimensional structure of a viral genome-delivery portal vertex. Nature Struc. Mol. Biol. 18, 597–604 (2011). This article describes the most recent of the three X-ray structures of portal protein rings (see also references 55 and 56).

    CAS  Article  Google Scholar 

  58. Zheng, H. et al. A conformational switch in bacteriophage P22 portal protein primes genome injection. Mol. Cell 29, 376–383 (2008).

    CAS  PubMed  PubMed Central  Article  Google Scholar 

  59. Cuervo, A., Vaney, M. C., Antson, A. A., Tavares, P. & Oliveira, L. Structural rearrangements between portal protein subunits are essential for viral DNA translocation. J. Biol. Chem. 282, 18907–18913 (2007).

    CAS  PubMed  Article  Google Scholar 

  60. Chen, D. H. et al. Structural basis for scaffolding-mediated assembly and maturation of a dsDNA virus. Proc. Natl Acad. Sci. USA 108, 1355–1360 (2011). This paper describes the highest-resolution asymmetrical three-dimensional reconstruction of procapsids that has been obtained to date by cryo-electron microscopy.

    CAS  PubMed  Article  PubMed Central  Google Scholar 

  61. Bazinet, C. & King, J. Initiation of P22 procapsid assembly in vivo. J. Mol. Biol. 202, 77–86 (1988).

    CAS  PubMed  Article  Google Scholar 

  62. Weigele, P., Sampson, L., Winn-Stapley, D. & Casjens, S. Molecular genetics of bacteriophage P22 scaffolding protein's functional domains. J. Mol. Biol. 348, 831–844 (2005).

    CAS  PubMed  Article  Google Scholar 

  63. Hendrix, R. W. Symmetry mismatch and DNA packaging in large bacteriophages. Proc. Natl Acad. Sci. USA 75, 4779–4783 (1978).

    CAS  PubMed  Article  PubMed Central  Google Scholar 

  64. Baumann, R. G., Mullaney, J. & Black, L. W. Portal fusion protein constraints on function in DNA packaging of bacteriophage T4. Mol. Microbiol. 61, 16–32 (2006).

    CAS  PubMed  Article  Google Scholar 

  65. Hugel, T. et al. Experimental test of connector rotation during DNA packaging into bacteriophage φ29 capsids. PLoS Biol. 5, e59 (2007).

    PubMed  PubMed Central  Article  CAS  Google Scholar 

  66. Ding, F. et al. Structure and assembly of the essential RNA ring component of a viral DNA packaging motor. Proc. Natl Acad. Sci. USA 108, 7357–7362 (2011).

    CAS  PubMed  Article  PubMed Central  Google Scholar 

  67. Reid, R., Zhang, F., Benson, S. & Anderson, D. Probing the structure of bacteriophage φ29 prohead RNA with specific mutations. J. Biol. Chem. 269, 18656–18661 (1994).

    CAS  PubMed  Google Scholar 

  68. Zhang, C., Tellinghuisen, T. & Guo, P. Confirmation of the helical structure of the 5′/3′ termini of the essential DNA packaging pRNA of phage φ29. RNA 1, 1041–1050 (1995).

    CAS  PubMed  PubMed Central  Google Scholar 

  69. Casjens, S. & Gilcrease, D. in Bacteriophages: Methods and Protocols (eds Clokie, M. & Kropinski, A.) 91–111 (Humana, Totowa, New Jersey, 2009).

    Book  Google Scholar 

  70. Jackson, E. N., Jackson, D. A. & Deans, R. J. EcoRI analysis of bacteriophage P22 DNA packaging. J. Mol. Biol. 118, 365–388 (1978).

    CAS  PubMed  Article  Google Scholar 

  71. Tye, B. K., Huberman, J. A. & Botstein, D. Non-random circular permutation of phage P22 DNA. J. Mol. Biol. 85, 501–528 (1974).

    CAS  PubMed  Article  Google Scholar 

  72. Adams, M. B., Hayden, M. & Casjens, S. On the sequential packaging of bacteriophage P22 DNA. J. Virol. 46, 673–677 (1983).

    CAS  PubMed  PubMed Central  Google Scholar 

  73. Meijer, W. J., Horcajadas, J. A. & Salas, M. φ29 family of phages. Microbiol. Mol. Biol. Rev. 65, 261–287 (2001).

    CAS  PubMed  PubMed Central  Article  Google Scholar 

  74. Mousset, S. & Thomas, R. Ter, a function which generates the ends of the mature λ chromosome. Nature 221, 242–245 (1969).

    CAS  PubMed  Article  Google Scholar 

  75. Poteete, A. R. & Botstein, D. Purification and properties of proteins essential to DNA encapsulation by phage P22. Virology 95, 565–573 (1979).

    CAS  PubMed  Article  Google Scholar 

  76. Al-Zahrani, A. S. et al. The small terminase, gp16, of bacteriophage T4 is a regulator of the DNA packaging motor. J. Biol. Chem. 284, 24490–24500 (2009).

    CAS  PubMed  PubMed Central  Article  Google Scholar 

  77. Nemecek, D. et al. Subunit conformations and assembly states of a DNA-translocating motor: the terminase of bacteriophage P22. J. Mol. Biol. 374, 817–836 (2007).

    CAS  PubMed  PubMed Central  Article  Google Scholar 

  78. Gual, A., Camacho, A. G. & Alonso, J. C. Functional analysis of the terminase large subunit, G2P, of Bacillus subtilis bacteriophage SPP1. J. Biol. Chem. 275, 35311–35319 (2000).

    CAS  PubMed  Article  Google Scholar 

  79. Leffers, G. & Rao, V. B. Biochemical characterization of an ATPase activity associated with the large packaging subunit gp17 from bacteriophage T4. J. Biol. Chem. 275, 37127–37136 (2000).

    CAS  PubMed  Article  Google Scholar 

  80. Maluf, N. K., Yang, Q. & Catalano, C. E. Self-association properties of the bacteriophage l terminase holoenzyme: implications for the DNA packaging motor. J. Mol. Biol. 347, 523–542 (2005).

    CAS  PubMed  Article  Google Scholar 

  81. Sun, S. et al. The structure of the phage T4 DNA packaging motor suggests a mechanism dependent on electrostatic forces. Cell 135, 1251–1262 (2008). This study uses cryo-electron microscopy to obtain a three-dimensionsal reconstruction of the phage T4 procapsid with the TerL ATPase bound to it.

    CAS  PubMed  Article  Google Scholar 

  82. Burroughs, A. M., Iyer, L. M. & Aravind, L. Comparative genomics and evolutionary trajectories of viral ATP dependent DNA-packaging systems. Genome Dyn. 3, 48–65 (2007).

    CAS  PubMed  Article  Google Scholar 

  83. Mitchell, M. S., Matsuzaki, S., Imai, S. & Rao, V. B. Sequence analysis of bacteriophage T4 DNA packaging/terminase genes 16 and 17 reveals a common ATPase center in the large subunit of viral terminases. Nucleic Acids Res. 30, 4009–4021 (2002).

    CAS  PubMed  PubMed Central  Article  Google Scholar 

  84. Guo, P., Peterson, C. & Anderson, D. Prohead and DNA-gp3-dependent ATPase activity of the DNA packaging protein gp16 of bacteriophage ϕ29. J. Mol. Biol. 197, 229–236 (1987).

    CAS  PubMed  Article  Google Scholar 

  85. Hwang, Y. & Feiss, M. Mutations affecting the high affinity ATPase center of gpA, the large subunit of bacteriophage l terminase, inactivate the endonuclease activity of terminase. J. Mol. Biol. 261, 524–535 (1996).

    CAS  PubMed  Article  Google Scholar 

  86. Morita, M., Tasaka, M. & Fujisawa, H. DNA packaging ATPase of bacteriophage T3. Virology 193, 748–752 (1993).

    CAS  PubMed  Article  Google Scholar 

  87. Oliveira, L., Henriques, A. O. & Tavares, P. Modulation of the viral ATPase activity by the portal protein correlates with DNA packaging efficiency. J. Biol. Chem. 281, 21914–21923 (2006).

    CAS  PubMed  Article  Google Scholar 

  88. Goetzinger, K. R. & Rao, V. B. Defining the ATPase center of bacteriophage T4 DNA packaging machine: requirement for a catalytic glutamate residue in the large terminase protein gp17. J. Mol. Biol. 331, 139–154 (2003).

    CAS  PubMed  Article  Google Scholar 

  89. Duffy, C. & Feiss, M. The large subunit of bacteriophage l's terminase plays a role in DNA translocation and packaging termination. J. Mol. Biol. 316, 547–561 (2002).

    CAS  PubMed  Article  Google Scholar 

  90. Kondabagil, K. R., Zhang, Z. & Rao, V. B. The DNA translocating ATPase of bacteriophage T4 packaging motor. J. Mol. Biol. 363, 786–799 (2006).

    CAS  PubMed  Article  Google Scholar 

  91. Tsay, J. M. et al. Mutations altering a structurally conserved loop-helix-loop region of a viral packaging motor change DNA translocation velocity and processivity. J. Biol. Chem. 285, 24282–24289 (2010).

    CAS  PubMed  PubMed Central  Article  Google Scholar 

  92. Tsay, J. M., Sippy, J., Feiss, M. & Smith, D. E. The Q motif of a viral packaging motor governs its force generation and communicates ATP recognition to DNA interaction. Proc. Natl Acad. Sci. USA 106, 14355–14360 (2009).

    CAS  PubMed  Article  PubMed Central  Google Scholar 

  93. Alam, T. I. et al. The headful packaging nuclease of bacteriophage T4. Mol. Microbiol. 69, 1180–1190 (2008).

    CAS  PubMed  Google Scholar 

  94. Smits, C. et al. Structural basis for the nuclease activity of a bacteriophage large terminase. EMBO Rep. 10, 592–598 (2009).

    CAS  PubMed  PubMed Central  Article  Google Scholar 

  95. Hwang, Y., Hang, J. Q., Neagle, J., Duffy, C. & Feiss, M. Endonuclease and helicase activities of bacteriophage l terminase: changing nearby residue 515 restores activity to the gpA K497D mutant enzyme. Virology 277, 204–214 (2000).

    CAS  PubMed  Article  Google Scholar 

  96. Zhang, Z. et al. A promiscuous DNA packaging machine from bacteriophage T4. PLoS Biol. 9, e1000592 (2011).

    CAS  PubMed  PubMed Central  Article  Google Scholar 

  97. Oliveira, L., Alonso, J. C. & Tavares, P. A defined in vitro system for DNA packaging by the bacteriophage SPP1: insights into the headful packaging mechanism. J. Mol. Biol. 353, 529–539 (2005).

    CAS  PubMed  Article  Google Scholar 

  98. de Beer, T. et al. Insights into specific DNA recognition during the assembly of a viral genome packaging machine. Mol. Cell 9, 981–991 (2002).

    CAS  PubMed  Article  Google Scholar 

  99. Hanagan, A., Meyer, J. D., Johnson, L., Manning, M. C. & Catalano, C. E. The phage lambda terminase enzyme: 2. Refolding of the gpNu1 subunit from the detergent-denatured and guanidinium hydrochloride-denatured state yields different oligomerization states and altered protein stabilities. Int. J. Biol. Macromol. 23, 37–48 (1998).

    CAS  PubMed  Article  Google Scholar 

  100. Nemecek, D., Lander, G. C., Johnson, J. E., Casjens, S. R. & Thomas, G. J. Jr. Assembly architecture and DNA binding of the bacteriophage P22 terminase small subunit. J. Mol. Biol. 383, 494–501 (2008).

    CAS  PubMed  PubMed Central  Article  Google Scholar 

  101. Zhao, H. et al. Crystal structure of the DNA-recognition component of the bacterial virus Sf6 genome-packaging machine. Proc. Natl Acad. Sci. USA 107, 1971–1976 (2010). This report presents the only currently known X-ray structure of TerS, the protein that is responsible for recognition of the DNA that will be packaged.

    CAS  PubMed  Article  PubMed Central  Google Scholar 

  102. Lin, H., Simon, M. N. & Black, L. W. Purification and characterization of the small subunit of phage T4 terminase, gp16, required for DNA packaging. J. Biol. Chem. 272, 3495–3501 (1997).

    CAS  PubMed  Article  Google Scholar 

  103. Chai, S., Lurz, R. & Alonso, J. C. The small subunit of the terminase enzyme of Bacillus subtilis bacteriophage SPP1 forms a specialized nucleoprotein complex with the packaging initiation region. J. Mol. Biol. 252, 386–398 (1995).

    CAS  PubMed  Article  Google Scholar 

  104. Casjens, S. R. & Thuman-Commike, P. A. Evolution of mosaically related tailed bacteriophage genomes seen through the lens of phage P22 virion assembly. Virology 411, 393–415 (2011).

    CAS  PubMed  Article  Google Scholar 

  105. Casjens, S. in Virus Structure and Assembly (ed. Casjens, S.) 75–148 (Jones and Bartlett, Boston, 1985).

    Google Scholar 

  106. Sun, S., Rao, V. B. & Rossmann, M. G. Genome packaging in viruses. Curr. Opin. Struct. Biol. 20, 114–120 (2010).

    CAS  PubMed  PubMed Central  Article  Google Scholar 

  107. Serwer, P. et al. DNA packaging-associated hyper-capsid expansion of bacteriophage T3. J. Mol. Biol. 397, 361–374 (2010).

    CAS  PubMed  PubMed Central  Article  Google Scholar 

  108. Yu, T. Y. & Schaefer, J. REDOR NMR characterization of DNA packaging in bacteriophage T4. J. Mol. Biol. 382, 1031–1042 (2008).

    CAS  PubMed  PubMed Central  Article  Google Scholar 

  109. Koti, J. S. et al. DNA packaging motor assembly intermediate of bacteriophage φ29. J. Mol. Biol. 381, 1114–1132 (2008).

    CAS  PubMed  PubMed Central  Article  Google Scholar 

  110. Maluf, N. K., Gaussier, H., Bogner, E., Feiss, M. & Catalano, C. E. Assembly of bacteriophage lambda terminase into a viral DNA maturation and packaging machine. Biochemistry 45, 15259–15268 (2006).

    CAS  PubMed  Article  Google Scholar 

  111. Chemla, Y. R. et al. Mechanism of force generation of a viral DNA packaging motor. Cell 122, 683–692 (2005).

    CAS  PubMed  Article  Google Scholar 

  112. Smith, D. et al. The bacteriophage φ29 portal motor can package DNA against a large internal force. Nature 413, 748–752 (2001). This important paper describes the first application of single-molecule optical tweezer technology to phage DNA packaging. It describes the basic rate and force properties of the phage ϕ29 motor.

    CAS  PubMed  Article  Google Scholar 

  113. Gope, R. & Serwer, P. Bacteriophage P22 in vitro DNA packaging monitored by agarose gel electrophoresis: rate of DNA entry into capsids. J. Virol. 47, 96–105 (1983).

    CAS  PubMed  PubMed Central  Google Scholar 

  114. Yang, Q., Catalano, C. E. & Maluf, N. K. Kinetic analysis of the genome packaging reaction in bacteriophage l. Biochemistry 48, 10705–10715 (2009).

    CAS  PubMed  Article  Google Scholar 

  115. Laemmli, U. K. & Favre, M. Maturation of the head of bacteriophage T4. I. DNA packaging events. J. Mol. Biol. 80, 575–599 (1973).

    CAS  PubMed  Article  Google Scholar 

  116. Fuller, D. N. et al. Measurements of single DNA molecule packaging dynamics in bacteriophage l reveal high forces, high motor processivity, and capsid transformations. J. Mol. Biol. 373, 1113–1122 (2007). This study defines the basic rate and force parameters of the phage λ packaging motor.

    CAS  PubMed  PubMed Central  Article  Google Scholar 

  117. Fuller, D. N., Raymer, D. M., Kottadiel, V. I., Rao, V. B. & Smith, D. E. Single phage T4 DNA packaging motors exhibit large force generation, high velocity, and dynamic variability. Proc. Natl Acad. Sci. USA 104, 16868–16873 (2007). This investigation defines the basic rate and force parameters of the phage T4 packaging motor.

    CAS  PubMed  Article  PubMed Central  Google Scholar 

  118. Rickgauer, J. P. et al. Portal motor velocity and internal force resisting viral DNA packaging in bacteriophage φ29. Biophys. J. 94, 159–167 (2008).

    CAS  PubMed  Article  Google Scholar 

  119. Moffitt, J. R. et al. Intersubunit coordination in a homomeric ring ATPase. Nature 457, 446–450 (2009). This important work refines optical tweezer analysis to be able to study the step size of DNA packaging (10 bp) and the coordination of the 2.5-bp substeps.

    CAS  PubMed  PubMed Central  Article  Google Scholar 

  120. Yu, J., Moffitt, J., Hetherington, C. L., Bustamante, C. & Oster, G. Mechanochemistry of a viral DNA packaging motor. J. Mol. Biol. 400, 186–203 (2010).

    CAS  PubMed  Article  Google Scholar 

  121. Draper, B. & Rao, V. B. An ATP hydrolysis sensor in the DNA packaging motor from bacteriophage T4 suggests an inchworm-type translocation mechanism. J. Mol. Biol. 369, 79–94 (2007).

    CAS  PubMed  Article  Google Scholar 

  122. Ray, K., Sabanayagam, C. R., Lakowicz, J. R. & Black, L. W. DNA crunching by a viral packaging motor: compression of a procapsid-portal stalled Y-DNA substrate. Virology 398, 224–232 (2010).

    CAS  PubMed  Article  Google Scholar 

  123. Ortega, M. E., Gaussier, H. & Catalano, C. E. The DNA maturation domain of gpA, the DNA packaging motor protein of bacteriophage lambda, contains an ATPase site associated with endonuclease activity. J. Mol. Biol. 373, 851–865 (2007).

    CAS  PubMed  PubMed Central  Article  Google Scholar 

  124. Ghosh-Kumar, M., Alam, T. I., Draper, B., Stack, J. D. & Rao, V. B. Regulation by interdomain communication of a headful packaging nuclease from bacteriophage T4. Nucleic Acids Res. 39, 2742–2755 (2011).

    CAS  PubMed  Article  Google Scholar 

  125. Yang, Q. & Catalano, C. E. A minimal kinetic model for a viral DNA packaging machine. Biochemistry 43, 289–299 (2004).

    CAS  PubMed  Article  Google Scholar 

  126. Gao, S. & Rao, V. B. Specificity of interactions among the DNA-packaging machine components of T4-related bacteriophages. J. Biol. Chem. 286, 3944–3956 (2011).

    CAS  PubMed  Article  Google Scholar 

  127. Woods, L., Terpening, C. & Catalano, C. E. Kinetic analysis of the endonuclease activity of phage l terminase: assembly of a catalytically competent nicking complex is rate-limiting. Biochemistry 36, 5777–5785 (1997).

    CAS  PubMed  Article  Google Scholar 

  128. Gaussier, H., Ortega, M. E., Maluf, N. K. & Catalano, C. E. Nucleotides regulate the conformational state of the small terminase subunit from bacteriophage lambda: implications for the assembly of a viral genome-packaging motor. Biochemistry 44, 9645–9656 (2005).

    CAS  PubMed  Article  Google Scholar 

  129. Aathavan, K. et al. Substrate interactions and promiscuity in a viral DNA packaging motor. Nature 461, 669–673 (2009).

    CAS  PubMed  PubMed Central  Article  Google Scholar 

  130. Oram, M., Sabanayagam, C. & Black, L. W. Modulation of the packaging reaction of bacteriophage T4 terminase by DNA structure. J. Mol. Biol. 381, 61–72 (2008).

    CAS  PubMed  PubMed Central  Article  Google Scholar 

  131. Leffers, G. & Rao, V. B. A discontinuous headful packaging model for packaging less than headful length DNA molecules by bacteriophage T4. J. Mol. Biol. 258, 839–850 (1996).

    CAS  PubMed  Article  Google Scholar 

  132. Xin, W. & Feiss, M. Function of IHF in l DNA packaging: I. Identification of the strong binding site for integration host factor and the locus for intrinsic bending in cosB. J. Mol. Biol. 230, 492–504 (1993).

    CAS  PubMed  Article  Google Scholar 

  133. Wu, H., Sampson, L., Parr, R. & Casjens, S. The DNA site utilized by bacteriophage P22 for initiation of DNA packaging. Mol. Microbiol. 45, 1631–1646 (2002).

    CAS  PubMed  Article  Google Scholar 

  134. Bukhari, A. I., Froshauer, S. & Botchan, M. Ends of bacteriophage Mu DNA. Nature 264, 580–583 (1976).

    CAS  PubMed  Article  Google Scholar 

  135. Groenen, M. A. & van de Putte, P. Mapping of a site for packaging of bacteriophage Mu DNA. Virology 144, 520–522 (1985).

    CAS  PubMed  Article  Google Scholar 

  136. Harel, J., Duplessis, L., Kahn, J. S. & DuBow, M. S. The cis-acting DNA sequences required in vivo for bacteriophage Mu helper-mediated transposition and packaging. Arch. Microbiol. 154, 67–72 (1990).

    CAS  PubMed  Article  Google Scholar 

  137. Casjens, S. et al. The chromosome of Shigella flexneri bacteriophage Sf6: complete nucleotide sequence, genetic mosaicism, and DNA packaging. J. Mol. Biol. 339, 379–394 (2004).

    CAS  PubMed  Article  Google Scholar 

  138. Casjens, S. R. et al. The generalized transducing Salmonella bacteriophage ES18: complete genome sequence and DNA packaging strategy. J. Bacteriol. 187, 1091–1104 (2005).

    CAS  PubMed  PubMed Central  Article  Google Scholar 

  139. Black, L. W. & Peng, G. Mechanistic coupling of bacteriophage T4 DNA packaging to components of the replication-dependent late transcription machinery. J. Biol. Chem. 281, 25635–25643 (2006).

    CAS  PubMed  Article  Google Scholar 

  140. Zhang, X. & Studier, F. W. Multiple roles of T7 RNA polymerase and T7 lysozyme during bacteriophage T7 infection. J. Mol. Biol. 340, 707–730 (2004).

    CAS  PubMed  Article  Google Scholar 

  141. Chung, Y. B., Nardone, C. & Hinkle, D. C. Bacteriophage T7 DNA packaging. III. A “hairpin” end formed on T7 concatemers may be an intermediate in the processing reaction. J. Mol. Biol. 216, 939–948 (1990).

    CAS  PubMed  Article  Google Scholar 

  142. Stewart, C. R. et al. The genome of Bacillus subtilis bacteriophage SPO1. J. Mol. Biol. 388, 48–70 (2009).

    CAS  PubMed  PubMed Central  Article  Google Scholar 

  143. Xin, W., Cai, Z. H. & Feiss, M. Function of IHF in l DNA packaging: II. Effects of mutations altering the IHF binding site and the intrinsic bend in cosB on l development. J. Mol. Biol. 230, 505–515 (1993).

    CAS  PubMed  Article  Google Scholar 

  144. Droge, A. & Tavares, P. in Viral Genome Packaging Machines: Genetics, Structure, and Mechanism (ed. Catalano, C.) 89–101 (Landes Bioscience, Georgetown, Texas, 2005).

    Book  Google Scholar 

  145. Feiss, M. & Catalano, C. in Viral Genome Packaging Machines: Genetics, Structure, and Mechanism (ed. Catalano, C.) 5–39 (Landes Bioscience, Georgetown, Texas, 2005).

    Book  Google Scholar 

  146. Frackman, S., Siegele, D. A. & Feiss, M. A functional domain of bacteriophage l terminase for prohead binding. J. Mol. Biol. 180, 283–300 (1984).

    CAS  PubMed  Article  Google Scholar 

  147. Morita, M., Tasaka, M. & Fujisawa, H. Structural and functional domains of the large subunit of the bacteriophage T3 DNA packaging enzyme: importance of the C-terminal region in prohead binding. J. Mol. Biol. 245, 635–644 (1995).

    CAS  PubMed  Article  Google Scholar 

  148. Oliveira, L., Cuervo, A. & Tavares, P. Direct interaction of the bacteriophage SPP1 packaging ATPase with the portal protein. J. Biol. Chem. 285, 7366–7373 (2010).

    CAS  PubMed  PubMed Central  Article  Google Scholar 

  149. Lin, H., Rao, V. B. & Black, L. W. Analysis of capsid portal protein and terminase functional domains: interaction sites required for DNA packaging in bacteriophage T4. J. Mol. Biol. 289, 249–260 (1999).

    CAS  PubMed  Article  Google Scholar 

  150. Casjens, S. et al. Bacteriophage P22 portal protein is part of the gauge that regulates packing density of intravirion DNA. J. Mol. Biol. 224, 1055–1074 (1992).

    CAS  PubMed  Article  Google Scholar 

  151. Isidro, A., Santos, M. A., Henriques, A. O. & Tavares, P. The high-resolution functional map of bacteriophage SPP1 portal protein. Mol. Microbiol. 51, 949–962 (2004).

    CAS  PubMed  Article  Google Scholar 

  152. Wieczorek, D. J. & Feiss, M. Defining cosQ, the site required for termination of bacteriophage l DNA packaging. Genetics 158, 495–506 (2001).

    CAS  PubMed  PubMed Central  Google Scholar 

  153. Cardarelli, L. et al. The crystal structure of bacteriophage HK97 gp6: defining a large family of head–tail connector proteins. J. Mol. Biol. 395, 754–768 (2010).

    CAS  PubMed  Article  Google Scholar 

  154. Lhuillier, S. et al. Structure of bacteriophage SPP1 head-to-tail connection reveals mechanism for viral DNA gating. Proc. Natl Acad. Sci. USA 106, 8507–8512 (2009).

    CAS  Article  PubMed  PubMed Central  Google Scholar 

  155. Olia, A. S. et al. Binding-induced stabilization and assembly of the phage P22 tail accessory factor gp4. J. Mol. Biol. 363, 558–576 (2006).

    CAS  PubMed  Article  Google Scholar 

  156. Olia, A. S., Bhardwaj, A., Joss, L., Casjens, S. & Cingolani, G. Role of gene 10 protein in the hierarchical assembly of the bacteriophage P22 portal vertex structure. Biochemistry 46, 8776–8784 (2007).

    CAS  PubMed  Article  Google Scholar 

  157. Olia, A. S., Casjens, S. & Cingolani, G. Structure of phage P22 cell envelope-penetrating needle. Nature Struct. Mol. Biol. 14, 1221–1227 (2007).

    CAS  Article  Google Scholar 

  158. Chattoraj, D. K. & Inman, R. B. Location of DNA ends in P2, 186, P4 and lambda bacteriophage heads. J. Mol. Biol. 87, 11–22 (1974).

    CAS  PubMed  Article  Google Scholar 

  159. Saigo, K. & Uchida, H. Connection of the right-hand terminus of DNA to the proximal end of the tail in bacteriophage lambda. Virology 61, 524–536 (1974).

    CAS  PubMed  Article  Google Scholar 

  160. Thomas, J. O., Sternberg, N. & Weisberg, R. Altered arrangement of the DNA in injection-defective lambda bacteriophage. J. Mol. Biol. 123, 149–161 (1978).

    CAS  PubMed  Article  Google Scholar 

  161. Plisson, C. et al. Structure of bacteriophage SPP1 tail reveals trigger for DNA ejection. EMBO J. 26, 3720–3728 (2007).

    CAS  PubMed  PubMed Central  Article  Google Scholar 

  162. Israel, V. E proteins of bacteriophage P22. I. Identification and ejection from wild-type and defective particles. J. Virol. 23, 91–97 (1977).

    CAS  PubMed  PubMed Central  Google Scholar 

  163. Wikoff, W. R. et al. Topologically linked protein rings in the bacteriophage HK97 capsid. Science 289, 2129–2133 (2000).

    CAS  PubMed  Article  Google Scholar 

  164. Gilcrease, E. B., Winn-Stapley, D. A., Hewitt, F. C., Joss, L. & Casjens, S. R. Nucleotide sequence of the head assembly gene cluster of bacteriophage L and decoration protein characterization. J. Bacteriol. 187, 2050–2057 (2005).

    CAS  PubMed  PubMed Central  Article  Google Scholar 

  165. Tang, L., Gilcrease, E. B., Casjens, S. R. & Johnson, J. E. Highly discriminatory binding of capsid-cementing proteins in bacteriophage L. Structure 14, 837–845 (2006).

    CAS  PubMed  Article  Google Scholar 

  166. Qin, L., Fokine, A., O'Donnell, E., Rao, V. B. & Rossmann, M. G. Structure of the small outer capsid protein, Soc: a clamp for stabilizing capsids of T4-like phages. J. Mol. Biol. 395, 728–741 (2010).

    CAS  PubMed  Article  Google Scholar 

  167. Sathaliyawala, T. et al. Functional analysis of the highly antigenic outer capsid protein, Hoc, a virus decoration protein from T4-like bacteriophages. Mol. Microbiol. 77, 444–455 (2010).

    CAS  PubMed  PubMed Central  Article  Google Scholar 

  168. Yang, Q., Maluf, N. K. & Catalano, C. E. Packaging of a unit-length viral genome: the role of nucleotides and the gpD decoration protein in stable nucleocapsid assembly in bacteriophage l. J. Mol. Biol. 383, 1037–1048 (2008).

    CAS  PubMed  Article  Google Scholar 

  169. Lee, T. J., Schwartz, C. & Guo, P. Construction of bacteriophage φ29 DNA packaging motor and its applications in nanotechnology and therapy. Ann. Biomed. Eng. 37, 2064–2081 (2009).

    PubMed  PubMed Central  Article  Google Scholar 

  170. Branton, D. et al. The potential and challenges of nanopore sequencing. Nature Biotech. 26, 1146–1153 (2008).

    CAS  Article  Google Scholar 

  171. Sun, S., Kondabagil, K., Gentz, P. M., Rossmann, M. G. & Rao, V. B. The structure of the ATPase that powers DNA packaging into bacteriophage T4 procapsids. Mol. Cell 25, 943–949 (2007).

    CAS  PubMed  Article  Google Scholar 

  172. Casjens, S. in Structural Biology of Viruses (eds Chiu, W., Burnett, R. & Garcea, R.) 3–37 (Oxford Univ. Press, Oxford, 1997).

    Google Scholar 

  173. Fuller, D. N. et al. Ionic effects on viral DNA packaging and portal motor function in bacteriophage ϕ29. Proc. Natl Acad. Sci. USA 104, 11245–11250 (2007). This paper describes the X-ray structure of phage T4 TerL, the ATPase that powers DNA packaging.

    CAS  PubMed  Article  PubMed Central  Google Scholar 

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Acknowledgements

I thank M. Feiss, V. Rao and S. Grimes for thoughtful reading of this manuscript before publication. My research is supported by the US National Institutes of Health grant RO1 AI074825.

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Glossary

Tailed bacteriophages

Double-stranded-DNA bacteriophages with a protein tail that attaches to a susceptible bacterium. DNA is injected into the host through the tail.

Prophages

Bacteriophage genomes that are physically integrated into the chromosome of the host bacterium.

Nanomotor

A molecular motor that functions on a nanometre scale to convert energy into directed physical movement of molecules.

Procapsid

The preformed protein container into which DNA is packaged during virion assembly.

Portal protein

The part of the DNA-packaging nanomotor that forms the hole or portal through the phage capsid; DNA enters and exits the virion through this portal.

Portal vertex

The unique vertex to which the portal protein and tails are attached. Icosahedral structures such as phage heads have 12 five-fold rotationally symmetrical vertices.

Low-angle X-ray scattering

A technique that is used to determine the radially averaged electron density of particles that make up an unoriented (as opposed to crystalline) sample. It is based on the deflection of a beam of X-rays away from its straight trajectory after it interacts with the particles in the sample.

Terminase

The DNA nanomotor protein complex that recognizes the DNA which will be packaged. The terminase contains the ATPase activity that converts chemical energy into mechanical movement, and also contains the active site for cleavage of double-stranded DNA (when such a site is present).

Concatemers

Long DNA molecules that contain multiple head-to-tail repeats of the phage genome sequence. Concatemers are often formed as the result of the DNA synthesis machinery replicating around a circular template multiple times without terminating.

Occam's razor

A line of reasoning that argues that the simplest explanation should be favoured until a more complex one is required by the observed data (attributed to the Franciscan friar Father William of Ockham).

Late-gene transcription factors

The phage-encoded early proteins (which are expressed early during infection) that cause transcription of the genes which are expressed at late times after infection.

IHF

(Integration host factor). An Escherichia coli protein that binds DNA and bends it at a sharp angle. IHF was first discovered as a cofactor of the phage λ enzyme integrase.

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Casjens, S. The DNA-packaging nanomotor of tailed bacteriophages. Nat Rev Microbiol 9, 647–657 (2011). https://doi.org/10.1038/nrmicro2632

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