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. 1993 Mar;64(3):824–835. doi: 10.1016/S0006-3495(93)81443-7

Nucleation and growth phases in the polymerization of coat and scaffolding subunits into icosahedral procapsid shells.

P E Prevelige Jr 1, D Thomas 1, J King 1
PMCID: PMC1262396  PMID: 8471727

Abstract

The polymerization of protein subunits into precursor shells empty of DNA is a critical process in the assembly of double-stranded DNA viruses. For the well-characterized icosahedral procapsid of phage P22, coat and scaffolding protein subunits do not assemble separately but, upon mixing, copolymerize into double-shelled procapsids in vitro. The polymerization reaction displays the characteristics of a nucleation limited reaction: a paucity of intermediate assembly states, a critical concentration, and kinetics displaying a lag phase. Partially formed shell intermediates were directly visualized during the growth phase by electron microscopy of the reaction mixture. The morphology of these intermediates suggests that assembly is a highly directed process. The initial rate of this reaction depends on the fifth power of the coat subunit concentration and the second or third power of the scaffolding concentration, suggesting that pentamer of coat protein and dimers or trimers of scaffolding protein, respectively, participate in the rate-limiting step.

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Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. Asakura S. A kinetic study of in vitro polymerization of flagellin. J Mol Biol. 1968 Jul 14;35(1):237–239. doi: 10.1016/s0022-2836(68)80051-8. [DOI] [PubMed] [Google Scholar]
  2. Baker T. S., Newcomb W. W., Booy F. P., Brown J. C., Steven A. C. Three-dimensional structures of maturable and abortive capsids of equine herpesvirus 1 from cryoelectron microscopy. J Virol. 1990 Feb;64(2):563–573. doi: 10.1128/jvi.64.2.563-573.1990. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Bazinet C., Benbasat J., King J., Carazo J. M., Carrascosa J. L. Purification and organization of the gene 1 portal protein required for phage P22 DNA packaging. Biochemistry. 1988 Mar 22;27(6):1849–1856. doi: 10.1021/bi00406a009. [DOI] [PubMed] [Google Scholar]
  4. Bazinet C., King J. Initiation of P22 procapsid assembly in vivo. J Mol Biol. 1988 Jul 5;202(1):77–86. doi: 10.1016/0022-2836(88)90520-7. [DOI] [PubMed] [Google Scholar]
  5. Bazinet C., Villafane R., King J. Novel second-site suppression of a cold-sensitive defect in phage P22 procapsid assembly. J Mol Biol. 1990 Dec 5;216(3):701–716. doi: 10.1016/0022-2836(90)90393-Z. [DOI] [PubMed] [Google Scholar]
  6. Botstein D., Waddell C. H., King J. Mechanism of head assembly and DNA encapsulation in Salmonella phage p22. I. Genes, proteins, structures and DNA maturation. J Mol Biol. 1973 Nov 15;80(4):669–695. doi: 10.1016/0022-2836(73)90204-0. [DOI] [PubMed] [Google Scholar]
  7. Butler P. J., Klug A. Assembly of the particle of tobacco mosaic virus from RNA and disks of protein. Nat New Biol. 1971 Jan 13;229(2):47–50. doi: 10.1038/newbio229047a0. [DOI] [PubMed] [Google Scholar]
  8. CASPAR D. L., KLUG A. Physical principles in the construction of regular viruses. Cold Spring Harb Symp Quant Biol. 1962;27:1–24. doi: 10.1101/sqb.1962.027.001.005. [DOI] [PubMed] [Google Scholar]
  9. Casjens S., King J. P22 morphogenesis. I: Catalytic scaffolding protein in capsid assembly. J Supramol Struct. 1974;2(2-4):202–224. doi: 10.1002/jss.400020215. [DOI] [PubMed] [Google Scholar]
  10. Casjens S. Molecular organization of the bacteriophage P22 coat protein shell. J Mol Biol. 1979 Jun 15;131(1):1–14. doi: 10.1016/0022-2836(79)90298-5. [DOI] [PubMed] [Google Scholar]
  11. Caspar D. L. Movement and self-control in protein assemblies. Quasi-equivalence revisited. Biophys J. 1980 Oct;32(1):103–138. doi: 10.1016/S0006-3495(80)84929-0. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Earnshaw W., Casjens S., Harrison S. C. Assembly of the head of bacteriophage P22: x-ray diffraction from heads, proheads and related structures. J Mol Biol. 1976 Jun 25;104(2):387–410. doi: 10.1016/0022-2836(76)90278-3. [DOI] [PubMed] [Google Scholar]
  13. Eppler K., Wyckoff E., Goates J., Parr R., Casjens S. Nucleotide sequence of the bacteriophage P22 genes required for DNA packaging. Virology. 1991 Aug;183(2):519–538. doi: 10.1016/0042-6822(91)90981-g. [DOI] [PubMed] [Google Scholar]
  14. Erickson H. P., Pantaloni D. The role of subunit entropy in cooperative assembly. Nucleation of microtubules and other two-dimensional polymers. Biophys J. 1981 May;34(2):293–309. doi: 10.1016/S0006-3495(81)84850-3. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Frieden C., Goddette D. W. Polymerization of actin and actin-like systems: evaluation of the time course of polymerization in relation to the mechanism. Biochemistry. 1983 Dec 6;22(25):5836–5843. doi: 10.1021/bi00294a023. [DOI] [PubMed] [Google Scholar]
  16. Fuller M. T., King J. Assembly in vitro of bacteriophage P22 procapsids from purified coat and scaffolding subunits. J Mol Biol. 1982 Apr 15;156(3):633–665. doi: 10.1016/0022-2836(82)90270-4. [DOI] [PubMed] [Google Scholar]
  17. Guo P. X., Erickson S., Xu W., Olson N., Baker T. S., Anderson D. Regulation of the phage phi 29 prohead shape and size by the portal vertex. Virology. 1991 Jul;183(1):366–373. doi: 10.1016/0042-6822(91)90149-6. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Kellenberger E. Form determination of the heads of bacteriophages. Eur J Biochem. 1990 Jun 20;190(2):233–248. doi: 10.1111/j.1432-1033.1990.tb15568.x. [DOI] [PubMed] [Google Scholar]
  19. King J., Casjens S. Catalytic head assembling protein in virus morphogenesis. Nature. 1974 Sep 13;251(5471):112–119. doi: 10.1038/251112a0. [DOI] [PubMed] [Google Scholar]
  20. Ladin B. F., Ihara S., Hampl H., Ben-Porat T. Pathway of assembly of herpesvirus capsids: an analysis using DNA+ temperature-sensitive mutants of pseudorabies virus. Virology. 1982 Jan 30;116(2):544–561. doi: 10.1016/0042-6822(82)90147-7. [DOI] [PubMed] [Google Scholar]
  21. Liddington R. C., Yan Y., Moulai J., Sahli R., Benjamin T. L., Harrison S. C. Structure of simian virus 40 at 3.8-A resolution. Nature. 1991 Nov 28;354(6351):278–284. doi: 10.1038/354278a0. [DOI] [PubMed] [Google Scholar]
  22. McKenna R., Xia D., Willingmann P., Ilag L. L., Krishnaswamy S., Rossmann M. G., Olson N. H., Baker T. S., Incardona N. L. Atomic structure of single-stranded DNA bacteriophage phi X174 and its functional implications. Nature. 1992 Jan 9;355(6356):137–143. doi: 10.1038/355137a0. [DOI] [PMC free article] [PubMed] [Google Scholar]
  23. Morin N., Boulanger P. Morphogenesis of human adenovirus type 2: sequence of entry of proteins into previral and viral particles. Virology. 1984 Jul 15;136(1):153–167. doi: 10.1016/0042-6822(84)90256-3. [DOI] [PubMed] [Google Scholar]
  24. Murialdo H., Becker A. Head morphogenesis of complex double-stranded deoxyribonucleic acid bacteriophages. Microbiol Rev. 1978 Sep;42(3):529–576. doi: 10.1128/mr.42.3.529-576.1978. [DOI] [PMC free article] [PubMed] [Google Scholar]
  25. Namba K., Stubbs G. Structure of tobacco mosaic virus at 3.6 A resolution: implications for assembly. Science. 1986 Mar 21;231(4744):1401–1406. doi: 10.1126/science.3952490. [DOI] [PubMed] [Google Scholar]
  26. Newcomb W. W., Brown J. C. Structure of the herpes simplex virus capsid: effects of extraction with guanidine hydrochloride and partial reconstitution of extracted capsids. J Virol. 1991 Feb;65(2):613–620. doi: 10.1128/jvi.65.2.613-620.1991. [DOI] [PMC free article] [PubMed] [Google Scholar]
  27. OOSAWA F., KASAI M. A theory of linear and helical aggregations of macromolecules. J Mol Biol. 1962 Jan;4:10–21. doi: 10.1016/s0022-2836(62)80112-0. [DOI] [PubMed] [Google Scholar]
  28. Preston V. G., Coates J. A., Rixon F. J. Identification and characterization of a herpes simplex virus gene product required for encapsidation of virus DNA. J Virol. 1983 Mar;45(3):1056–1064. doi: 10.1128/jvi.45.3.1056-1064.1983. [DOI] [PMC free article] [PubMed] [Google Scholar]
  29. Prevelige P. E., Jr, Thomas D., King J. Scaffolding protein regulates the polymerization of P22 coat subunits into icosahedral shells in vitro. J Mol Biol. 1988 Aug 20;202(4):743–757. doi: 10.1016/0022-2836(88)90555-4. [DOI] [PubMed] [Google Scholar]
  30. Raghavendra K., Kelly J. A., Khairallah L., Schuster T. M. Structure and function of disk aggregates of the coat protein of tobacco mosaic virus. Biochemistry. 1988 Oct 4;27(20):7583–7588. doi: 10.1021/bi00420a002. [DOI] [PubMed] [Google Scholar]
  31. Raghavendra K., Salunke D. M., Caspar D. L., Schuster T. M. Disk aggregates of tobacco mosaic virus protein in solution: electron microscopy observations. Biochemistry. 1986 Oct 7;25(20):6276–6279. doi: 10.1021/bi00368a066. [DOI] [PubMed] [Google Scholar]
  32. Rayment I., Baker T. S., Caspar D. L., Murakami W. T. Polyoma virus capsid structure at 22.5 A resolution. Nature. 1982 Jan 14;295(5845):110–115. doi: 10.1038/295110a0. [DOI] [PMC free article] [PubMed] [Google Scholar]
  33. Rossmann M. G. Constraints on the assembly of spherical virus particles. Virology. 1984 Apr 15;134(1):1–11. doi: 10.1016/0042-6822(84)90267-8. [DOI] [PubMed] [Google Scholar]
  34. Rossmann M. G., Johnson J. E. Icosahedral RNA virus structure. Annu Rev Biochem. 1989;58:533–573. doi: 10.1146/annurev.bi.58.070189.002533. [DOI] [PubMed] [Google Scholar]
  35. Rueckert R. R., Dunker A. K., Stoltzfus C. M. The structure of mouse-Elberfeld virus: a model. Proc Natl Acad Sci U S A. 1969 Mar;62(3):912–919. doi: 10.1073/pnas.62.3.912. [DOI] [PMC free article] [PubMed] [Google Scholar]
  36. Salunke D. M., Caspar D. L., Garcea R. L. Self-assembly of purified polyomavirus capsid protein VP1. Cell. 1986 Sep 12;46(6):895–904. doi: 10.1016/0092-8674(86)90071-1. [DOI] [PubMed] [Google Scholar]
  37. Showe M. K., Black L. W. Assembly core of bacteriophage T4: an intermediate in head formation. Nat New Biol. 1973 Mar 21;242(116):70–75. doi: 10.1038/newbio242070a0. [DOI] [PubMed] [Google Scholar]
  38. Silva A. M., Rossmann M. G. Refined structure of southern bean mosaic virus at 2.9 A resolution. J Mol Biol. 1987 Sep 5;197(1):69–87. doi: 10.1016/0022-2836(87)90610-3. [DOI] [PubMed] [Google Scholar]
  39. Sorger P. K., Stockley P. G., Harrison S. C. Structure and assembly of turnip crinkle virus. II. Mechanism of reassembly in vitro. J Mol Biol. 1986 Oct 20;191(4):639–658. doi: 10.1016/0022-2836(86)90451-1. [DOI] [PubMed] [Google Scholar]
  40. Steven A. C., Bauer A. C., Bisher M. E., Robey F. A., Black L. W. The maturation-dependent conformational change of phage T4 capsid involves the translocation of specific epitopes between the inner and the outer capsid surfaces. J Struct Biol. 1991 Jun;106(3):221–236. doi: 10.1016/1047-8477(91)90072-5. [DOI] [PubMed] [Google Scholar]
  41. Steven A. C., Greenstone H., Bauer A. C., Williams R. W. The maturation-dependent conformational change of the major capsid protein of bacteriophage T4 involves a substantial change in secondary structure. Biochemistry. 1990 Jun 12;29(23):5556–5561. doi: 10.1021/bi00475a020. [DOI] [PubMed] [Google Scholar]
  42. Thomas D., Prevelige P., Jr A pilot protein participates in the initiation of P22 procapsid assembly. Virology. 1991 Jun;182(2):673–681. doi: 10.1016/0042-6822(91)90608-e. [DOI] [PubMed] [Google Scholar]
  43. Tobacman L. S., Korn E. D. The kinetics of actin nucleation and polymerization. J Biol Chem. 1983 Mar 10;258(5):3207–3214. [PubMed] [Google Scholar]
  44. Tsao J., Chapman M. S., Agbandje M., Keller W., Smith K., Wu H., Luo M., Smith T. J., Rossmann M. G., Compans R. W. The three-dimensional structure of canine parvovirus and its functional implications. Science. 1991 Mar 22;251(5000):1456–1464. doi: 10.1126/science.2006420. [DOI] [PubMed] [Google Scholar]
  45. Voter W. A., Erickson H. P. The kinetics of microtubule assembly. Evidence for a two-stage nucleation mechanism. J Biol Chem. 1984 Aug 25;259(16):10430–10438. [PubMed] [Google Scholar]
  46. Wegner A., Engel J. Kinetics of the cooperative association of actin to actin filaments. Biophys Chem. 1975 Jul;3(3):215–225. doi: 10.1016/0301-4622(75)80013-5. [DOI] [PubMed] [Google Scholar]

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