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. 1997 Mar;71(3):2211–2216. doi: 10.1128/jvi.71.3.2211-2216.1997

Two proline residues are essential in the calcium-binding activity of rotavirus VP7 outer capsid protein.

R Gajardo 1, P Vende 1, D Poncet 1, J Cohen 1
PMCID: PMC191328  PMID: 9032355

Abstract

Rotavirus maturation and stability of the outer capsid are calcium-dependent processes. It has been shown previously that the concentration of Ca2+-solubilizing outer capsid proteins from rotavirus particles is dependent on the virus strain. This property of viral particles has been associated with the gene coding for VP7 (gene 9). In this study the correlation between VP7 and resistance to low [Ca2+] was confirmed by analyzing the origin of gene 9 from reassortant viruses prepared under the selective pressure of low [Ca2+]. After chemical mutagenesis, we selected mutant viruses of the bovine strain RF that are more resistant to low [Ca2+]. The genes coding for the VP7 proteins of these independent mutants have been sequenced. Sequence analysis confirmed that these mutants are independent and revealed that all mutant VP7 proteins have proline 75 changed to leucine and have an outer capsid that solubilized at low [Ca2+]. The mutation of proline 279 to serine is found in all but two mutants. The phenotype of mutants having a single proline change can be distinguished from the phenotype of mutants having two proline changes. Sequence analysis showed that position 75 is in a region (amino acids 65 to 78) of great variability and that proline 75 is present in most of the bovine strains. In contrast, proline 279 is in a conserved region and is conserved in all the VP7 sequences in data banks. This region is rich in oxygenated residues that are correctly allocated in the metal-coordinating positions of the Ca2+-binding EF-hand structure pattern, suggesting that this region is important in the Ca2+ binding of VP7.

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Selected References

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  1. Burmeister W. P., Ruigrok R. W., Cusack S. The 2.2 A resolution crystal structure of influenza B neuraminidase and its complex with sialic acid. EMBO J. 1992 Jan;11(1):49–56. doi: 10.1002/j.1460-2075.1992.tb05026.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Chen D. Y., Estes M. K., Ramig R. F. Specific interactions between rotavirus outer capsid proteins VP4 and VP7 determine expression of a cross-reactive, neutralizing VP4-specific epitope. J Virol. 1992 Jan;66(1):432–439. doi: 10.1128/jvi.66.1.432-439.1992. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Cloyd M. W., Lynn W. S., Ramsey K., Baron S. Inhibition of human immunodeficiency virus (HIV-1) infection by diphenylhydantoin (dilantin) implicates role of cellular calcium in virus life cycle. Virology. 1989 Dec;173(2):581–590. doi: 10.1016/0042-6822(89)90569-2. [DOI] [PubMed] [Google Scholar]
  4. Cohen J., Laporte J., Charpilienne A., Scherrer R. Activation of rotavirus RNA polymerase by calcium chelation. Arch Virol. 1979;60(3-4):177–186. doi: 10.1007/BF01317489. [DOI] [PubMed] [Google Scholar]
  5. Concha N. O., Head J. F., Kaetzel M. A., Dedman J. R., Seaton B. A. Rat annexin V crystal structure: Ca(2+)-induced conformational changes. Science. 1993 Sep 3;261(5126):1321–1324. doi: 10.1126/science.8362244. [DOI] [PubMed] [Google Scholar]
  6. Devereux J., Haeberli P., Smithies O. A comprehensive set of sequence analysis programs for the VAX. Nucleic Acids Res. 1984 Jan 11;12(1 Pt 1):387–395. doi: 10.1093/nar/12.1part1.387. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Dormitzer P. R., Greenberg H. B. Calcium chelation induces a conformational change in recombinant herpes simplex virus-1-expressed rotavirus VP7. Virology. 1992 Aug;189(2):828–832. doi: 10.1016/0042-6822(92)90616-w. [DOI] [PubMed] [Google Scholar]
  8. Durham A. C., Haidar M. A. Cation binding by tobacco rattle virus. Virology. 1977 Apr;77(2):520–523. doi: 10.1016/0042-6822(77)90477-9. [DOI] [PubMed] [Google Scholar]
  9. Durham A. C., Hendry D. A. Cation binding by tobacco mosaic virus. Virology. 1977 Apr;77(2):510–519. doi: 10.1016/0042-6822(77)90476-7. [DOI] [PubMed] [Google Scholar]
  10. Ebenbichler C. F., Stoiber H., Schneider R., Patsch J. R., Dierich M. P. The human immunodeficiency virus type 1 transmembrane gp41 protein is a calcium-binding protein and interacts with the putative second-receptor molecules in a calcium-dependent manner. J Virol. 1996 Mar;70(3):1723–1728. doi: 10.1128/jvi.70.3.1723-1728.1996. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Estes M. K., Cohen J. Rotavirus gene structure and function. Microbiol Rev. 1989 Dec;53(4):410–449. doi: 10.1128/mr.53.4.410-449.1989. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Haynes J. I., 2nd, Chang D., Consigli R. A. Mutations in the putative calcium-binding domain of polyomavirus VP1 affect capsid assembly. J Virol. 1993 May;67(5):2486–2495. doi: 10.1128/jvi.67.5.2486-2495.1993. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Irurzun A., Arroyo J., Alvarez A., Carrasco L. Enhanced intracellular calcium concentration during poliovirus infection. J Virol. 1995 Aug;69(8):5142–5146. doi: 10.1128/jvi.69.8.5142-5146.1995. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Jones T. A., Liljas L. Structure of satellite tobacco necrosis virus after crystallographic refinement at 2.5 A resolution. J Mol Biol. 1984 Aug 25;177(4):735–767. doi: 10.1016/0022-2836(84)90047-0. [DOI] [PubMed] [Google Scholar]
  15. Kawasaki H., Kretsinger R. H. Calcium-binding proteins. 1: EF-hands. Protein Profile. 1994;1(4):343–517. [PubMed] [Google Scholar]
  16. Laakso M. M., Heaton L. A. Asp-->Asn substitutions in the putative calcium-binding site of the turnip crinkle virus coat protein affect virus movement in plants. Virology. 1993 Dec;197(2):774–777. doi: 10.1006/viro.1993.1655. [DOI] [PubMed] [Google Scholar]
  17. Lazdins I., Coulson B. S., Kirkwood C., Dyall-Smith M., Masendycz P. J., Sonza S., Holmes I. H. Rotavirus antigenicity is affected by the genetic context and glycosylation of VP7. Virology. 1995 May 10;209(1):80–89. doi: 10.1006/viro.1995.1232. [DOI] [PubMed] [Google Scholar]
  18. 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]
  19. Ludlow J. W., Consigli R. A. Localization of calcium on the polyomavirus VP1 capsid protein. J Virol. 1987 Sep;61(9):2934–2937. doi: 10.1128/jvi.61.9.2934-2937.1987. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Maass D. R., Atkinson P. H. Rotavirus proteins VP7, NS28, and VP4 form oligomeric structures. J Virol. 1990 Jun;64(6):2632–2641. doi: 10.1128/jvi.64.6.2632-2641.1990. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. McPhalen C. A., Strynadka N. C., James M. N. Calcium-binding sites in proteins: a structural perspective. Adv Protein Chem. 1991;42:77–144. doi: 10.1016/s0065-3233(08)60535-5. [DOI] [PubMed] [Google Scholar]
  22. Michelangeli F., Liprandi F., Chemello M. E., Ciarlet M., Ruiz M. C. Selective depletion of stored calcium by thapsigargin blocks rotavirus maturation but not the cytopathic effect. J Virol. 1995 Jun;69(6):3838–3847. doi: 10.1128/jvi.69.6.3838-3847.1995. [DOI] [PMC free article] [PubMed] [Google Scholar]
  23. Michelangeli F., Ruiz M. C., del Castillo J. R., Ludert J. E., Liprandi F. Effect of rotavirus infection on intracellular calcium homeostasis in cultured cells. Virology. 1991 Apr;181(2):520–527. doi: 10.1016/0042-6822(91)90884-e. [DOI] [PubMed] [Google Scholar]
  24. Nokta M., Eaton D., Steinsland O. S., Albrecht T. Ca2+ responses in cytomegalovirus-infected fibroblasts of human origin. Virology. 1987 Apr;157(2):259–267. doi: 10.1016/0042-6822(87)90268-6. [DOI] [PubMed] [Google Scholar]
  25. Pattanayek R., Stubbs G. Structure of the U2 strain of tobacco mosaic virus refined at 3.5 A resolution using X-ray fiber diffraction. J Mol Biol. 1992 Nov 20;228(2):516–528. doi: 10.1016/0022-2836(92)90839-c. [DOI] [PubMed] [Google Scholar]
  26. Poncet D., Corthier G., Charpilienne A., Cohen J. A recombinant vaccinia virus expressing the major capsid protein of Simian rotavirus-induced anti-rotavirus antibodies. Virus Res. 1990 Mar;15(3):267–274. doi: 10.1016/0168-1702(90)90034-9. [DOI] [PubMed] [Google Scholar]
  27. Poruchynsky M. S., Maass D. R., Atkinson P. H. Calcium depletion blocks the maturation of rotavirus by altering the oligomerization of virus-encoded proteins in the ER. J Cell Biol. 1991 Aug;114(4):651–656. doi: 10.1083/jcb.114.4.651. [DOI] [PMC free article] [PubMed] [Google Scholar]
  28. Ramig R. F. Isolation and genetic characterization of temperature-sensitive mutants of simian rotavirus SA11. Virology. 1982 Jul 15;120(1):93–105. doi: 10.1016/0042-6822(82)90009-5. [DOI] [PubMed] [Google Scholar]
  29. Ramig R. F. Isolation and genetic characterization of temperature-sensitive mutants that define five additional recombination groups in simian rotavirus SA11. Virology. 1983 Oct 30;130(2):464–473. doi: 10.1016/0042-6822(83)90100-9. [DOI] [PubMed] [Google Scholar]
  30. Ruiz M. C., Charpilienne A., Liprandi F., Gajardo R., Michelangeli F., Cohen J. The concentration of Ca2+ that solubilizes outer capsid proteins from rotavirus particles is dependent on the strain. J Virol. 1996 Aug;70(8):4877–4883. doi: 10.1128/jvi.70.8.4877-4883.1996. [DOI] [PMC free article] [PubMed] [Google Scholar]
  31. Shahrabadi M. S., Lee P. W. Bovine rotavirus maturation is a calcium-dependent process. Virology. 1986 Jul 30;152(2):298–307. doi: 10.1016/0042-6822(86)90133-9. [DOI] [PubMed] [Google Scholar]
  32. Tsien R., Pozzan T. Measurement of cytosolic free Ca2+ with quin2. Methods Enzymol. 1989;172:230–262. doi: 10.1016/s0076-6879(89)72017-6. [DOI] [PubMed] [Google Scholar]
  33. Wery J. P., Reddy V. S., Hosur M. V., Johnson J. E. The refined three-dimensional structure of an insect virus at 2.8 A resolution. J Mol Biol. 1994 Jan 14;235(2):565–586. doi: 10.1006/jmbi.1994.1014. [DOI] [PubMed] [Google Scholar]
  34. Xu Z., Woode G. N. Studies on the influence of the VP7 gene on rotavirus replication. Virology. 1994 Jan;198(1):394–398. doi: 10.1006/viro.1994.1049. [DOI] [PubMed] [Google Scholar]
  35. Yamashita M. M., Wesson L., Eisenman G., Eisenberg D. Where metal ions bind in proteins. Proc Natl Acad Sci U S A. 1990 Aug;87(15):5648–5652. doi: 10.1073/pnas.87.15.5648. [DOI] [PMC free article] [PubMed] [Google Scholar]

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