Skip to main content
Journal of Virology logoLink to Journal of Virology
. 1995 Nov;69(11):7264–7268. doi: 10.1128/jvi.69.11.7264-7268.1995

Active foamy virus proteinase is essential for virus infectivity but not for formation of a Pol polyprotein.

J Konvalinka 1, M Löchelt 1, H Zentgraf 1, R M Flügel 1, H G Kräusslich 1
PMCID: PMC189650  PMID: 7474150

Abstract

To analyze proteolytic processing of foamy (spuma) retroviruses, two mutations were generated in the presumed active-site triplet Asp-Ser-Gly in the predicted proteinase (PR) region of the human foamy virus (HSRV). The mutations changed either the presumed catalytic aspartic acid residue to a catalytically incompetent alanine or the adjacent serine to a threonine found in most cellular and retroviral proteases at this position. Both mutations were cloned into the full-length infectious HSRV DNA clone. Wild-type and S/T mutant genomes directed the synthesis of particles with similar infectious titers, while the HSRV D/A PR mutant was noninfectious. Immunoblot analysis of transfected cells revealed identical patterns for the wild-type and for the S/T PR mutant. HSRV D/A mutant-transfected cells expressed only a single Gag polyprotein of 78 kDa instead of the 78-kDa-74-kDa doublet found in HSRV-infected or wild-type-transfected cells. Analysis with pol-specific antisera yielded a protein of approximately 120 kDa reactive with antisera against pol- but not gag-specific domains. No Gag-Pol polyprotein was detected in this study. Electron microscopy analysis of transfected cells showed heterogeneous particle morphology in the case of the D/A mutant, with particles of normal appearance and particles of aberrant size and shape. These results indicate that foamy viruses have an aspartic PR that is essential for infectivity but not for formation of the 120-kDa Pol polyprotein.

Full Text

The Full Text of this article is available as a PDF (1.3 MB).

Selected References

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

  1. Achong B. G., Mansell P. W., Epstein M. A., Clifford P. An unusual virus in cultures from a human nasopharyngeal carcinoma. J Natl Cancer Inst. 1971 Feb;46(2):299–307. [PubMed] [Google Scholar]
  2. Aguzzi A., Wagner E. F., Netzer K. O., Bothe K., Anhauser I., Rethwilm A. Human foamy virus proteins accumulate in neurons and induce multinucleated giant cells in the brain of transgenic mice. Am J Pathol. 1993 Apr;142(4):1061–1071. [PMC free article] [PubMed] [Google Scholar]
  3. Bartholomä A., Muranyi W., Flügel R. M. Bacterial expression of the capsid antigen domain and identification of native gag proteins in spumavirus-infected cells. Virus Res. 1992 Apr;23(1-2):27–38. doi: 10.1016/0168-1702(92)90065-h. [DOI] [PubMed] [Google Scholar]
  4. Benzair A. B., Rhodes-Feuillette A., Emanoïl-Ravicovitch R., Peries J. Reverse transcriptase from simian foamy virus serotype 1: purification and characterization. J Virol. 1982 Nov;44(2):720–724. doi: 10.1128/jvi.44.2.720-724.1982. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Carrière C., Gay B., Chazal N., Morin N., Boulanger P. Sequence requirements for encapsidation of deletion mutants and chimeras of human immunodeficiency virus type 1 Gag precursor into retrovirus-like particles. J Virol. 1995 Apr;69(4):2366–2377. doi: 10.1128/jvi.69.4.2366-2377.1995. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Davies D. R. The structure and function of the aspartic proteinases. Annu Rev Biophys Biophys Chem. 1990;19:189–215. doi: 10.1146/annurev.bb.19.060190.001201. [DOI] [PubMed] [Google Scholar]
  7. Kaplan A. H., Zack J. A., Knigge M., Paul D. A., Kempf D. J., Norbeck D. W., Swanstrom R. Partial inhibition of the human immunodeficiency virus type 1 protease results in aberrant virus assembly and the formation of noninfectious particles. J Virol. 1993 Jul;67(7):4050–4055. doi: 10.1128/jvi.67.7.4050-4055.1993. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Katz R. A., Skalka A. M. The retroviral enzymes. Annu Rev Biochem. 1994;63:133–173. doi: 10.1146/annurev.bi.63.070194.001025. [DOI] [PubMed] [Google Scholar]
  9. Kirchner J., Sandmeyer S. Proteolytic processing of Ty3 proteins is required for transposition. J Virol. 1993 Jan;67(1):19–28. doi: 10.1128/jvi.67.1.19-28.1993. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Kohl N. E., Emini E. A., Schleif W. A., Davis L. J., Heimbach J. C., Dixon R. A., Scolnick E. M., Sigal I. S. Active human immunodeficiency virus protease is required for viral infectivity. Proc Natl Acad Sci U S A. 1988 Jul;85(13):4686–4690. doi: 10.1073/pnas.85.13.4686. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Konvalinka J., Litterst M. A., Welker R., Kottler H., Rippmann F., Heuser A. M., Kräusslich H. G. An active-site mutation in the human immunodeficiency virus type 1 proteinase (PR) causes reduced PR activity and loss of PR-mediated cytotoxicity without apparent effect on virus maturation and infectivity. J Virol. 1995 Nov;69(11):7180–7186. doi: 10.1128/jvi.69.11.7180-7186.1995. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Kräusslich H. G., Wimmer E. Viral proteinases. Annu Rev Biochem. 1988;57:701–754. doi: 10.1146/annurev.bi.57.070188.003413. [DOI] [PubMed] [Google Scholar]
  13. Kunkel T. A. Rapid and efficient site-specific mutagenesis without phenotypic selection. Proc Natl Acad Sci U S A. 1985 Jan;82(2):488–492. doi: 10.1073/pnas.82.2.488. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Löchelt M., Yu S. F., Linial M. L., Flügel R. M. The human foamy virus internal promoter is required for efficient gene expression and infectivity. Virology. 1995 Jan 10;206(1):601–610. doi: 10.1016/s0042-6822(95)80077-8. [DOI] [PubMed] [Google Scholar]
  15. Löchelt M., Zentgraf H., Flügel R. M. Construction of an infectious DNA clone of the full-length human spumaretrovirus genome and mutagenesis of the bel 1 gene. Virology. 1991 Sep;184(1):43–54. doi: 10.1016/0042-6822(91)90820-2. [DOI] [PubMed] [Google Scholar]
  16. Mergener K., Fäcke M., Welker R., Brinkmann V., Gelderblom H. R., Kräusslich H. G. Analysis of HIV particle formation using transient expression of subviral constructs in mammalian cells. Virology. 1992 Jan;186(1):25–39. doi: 10.1016/0042-6822(92)90058-w. [DOI] [PubMed] [Google Scholar]
  17. Netzer K. O., Rethwilm A., Maurer B., ter Meulen V. Identification of the major immunogenic structural proteins of human foamy virus. J Gen Virol. 1990 May;71(Pt 5):1237–1241. doi: 10.1099/0022-1317-71-5-1237. [DOI] [PubMed] [Google Scholar]
  18. Netzer K. O., Schliephake A., Maurer B., Watanabe R., Aguzzi A., Rethwilm A. Identification of pol-related gene products of human foamy virus. Virology. 1993 Jan;192(1):336–338. doi: 10.1006/viro.1993.1039. [DOI] [PubMed] [Google Scholar]
  19. Pahl A., Flügel R. M. Endonucleolytic cleavages and DNA-joining activities of the integration protein of human foamy virus. J Virol. 1993 Sep;67(9):5426–5434. doi: 10.1128/jvi.67.9.5426-5434.1993. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Pearl L. H., Taylor W. R. A structural model for the retroviral proteases. Nature. 1987 Sep 24;329(6137):351–354. doi: 10.1038/329351a0. [DOI] [PubMed] [Google Scholar]
  21. Peng C., Ho B. K., Chang T. W., Chang N. T. Role of human immunodeficiency virus type 1-specific protease in core protein maturation and viral infectivity. J Virol. 1989 Jun;63(6):2550–2556. doi: 10.1128/jvi.63.6.2550-2556.1989. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Roberts N. A., Martin J. A., Kinchington D., Broadhurst A. V., Craig J. C., Duncan I. B., Galpin S. A., Handa B. K., Kay J., Kröhn A. Rational design of peptide-based HIV proteinase inhibitors. Science. 1990 Apr 20;248(4953):358–361. doi: 10.1126/science.2183354. [DOI] [PubMed] [Google Scholar]
  23. Tanese N., Goff S. P. Domain structure of the Moloney murine leukemia virus reverse transcriptase: mutational analysis and separate expression of the DNA polymerase and RNase H activities. Proc Natl Acad Sci U S A. 1988 Mar;85(6):1777–1781. doi: 10.1073/pnas.85.6.1777. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. Verma I. M. Studies on reverse transcriptase of RNA tumor viruses III. Properties of purified Moloney murine leukemia virus DNA polymerase and associated RNase H. J Virol. 1975 Apr;15(4):843–854. doi: 10.1128/jvi.15.4.843-854.1975. [DOI] [PMC free article] [PubMed] [Google Scholar]
  25. Wlodawer A., Erickson J. W. Structure-based inhibitors of HIV-1 protease. Annu Rev Biochem. 1993;62:543–585. doi: 10.1146/annurev.bi.62.070193.002551. [DOI] [PubMed] [Google Scholar]
  26. Wlodawer A., Miller M., Jaskólski M., Sathyanarayana B. K., Baldwin E., Weber I. T., Selk L. M., Clawson L., Schneider J., Kent S. B. Conserved folding in retroviral proteases: crystal structure of a synthetic HIV-1 protease. Science. 1989 Aug 11;245(4918):616–621. doi: 10.1126/science.2548279. [DOI] [PubMed] [Google Scholar]
  27. Yu S. F., Linial M. L. Analysis of the role of the bel and bet open reading frames of human foamy virus by using a new quantitative assay. J Virol. 1993 Nov;67(11):6618–6624. doi: 10.1128/jvi.67.11.6618-6624.1993. [DOI] [PMC free article] [PubMed] [Google Scholar]

Articles from Journal of Virology are provided here courtesy of American Society for Microbiology (ASM)

RESOURCES