Domains upstream of the protease (PR) in human immunodeficiency virus type 1 Gag-Pol influence PR autoprocessing (original) (raw)

Abstract

A critical step in the formation of infectious retroviral particles is the activation of the virally encoded protease (PR) and its release from the Gag-Pol precursor polyprotein. To identify factors that influence this step, the maturation of human immunodeficiency virus type 1 PR from various Gag-PR polyproteins was assayed in vitro by a using rabbit reticulocyte lysate as a coupled transcription-translation-autoprocessing system. Highly efficient autoprocessing was detected with polyproteins containing the viral nucleocapsid (NC) domain. In contrast, polyproteins consisting of only p6 and PR domains or containing a truncated NC domain exhibited no autoprocessing activity. Experiments designed to test the dimerization capability of short PR polyproteins revealed that precursors containing the NC domain exhibited very efficient homotypic protein-protein interactions while PR precursors consisting of only p6 and PR did not interact efficiently. The strong correlation between autoprocessing activity and PR polyprotein precursor dimerization suggests that NC and p6* domains play a role in PR activation by influencing the dimerization of the PR domain in the precursor.

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Selected References

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  1. Bennett R. P., Nelle T. D., Wills J. W. Functional chimeras of the Rous sarcoma virus and human immunodeficiency virus gag proteins. J Virol. 1993 Nov;67(11):6487–6498. doi: 10.1128/jvi.67.11.6487-6498.1993. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Cameron C. E., Grinde B., Jentoft J., Leis J., Weber I. T., Copeland T. D., Wlodawer A. Mechanism of inhibition of the retroviral protease by a Rous sarcoma virus peptide substrate representing the cleavage site between the gag p2 and p10 proteins. J Biol Chem. 1992 Nov 25;267(33):23735–23741. [PubMed] [Google Scholar]
  3. Carter C., Zybarth G. Processing of retroviral Gag polyproteins: an in vitro approach. Methods Enzymol. 1994;241:227–253. doi: 10.1016/0076-6879(94)41067-4. [DOI] [PubMed] [Google Scholar]
  4. Chien C. T., Bartel P. L., Sternglanz R., Fields S. The two-hybrid system: a method to identify and clone genes for proteins that interact with a protein of interest. Proc Natl Acad Sci U S A. 1991 Nov 1;88(21):9578–9582. doi: 10.1073/pnas.88.21.9578. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Darlix J. L., Gabus C., Nugeyre M. T., Clavel F., Barré-Sinoussi F. Cis elements and trans-acting factors involved in the RNA dimerization of the human immunodeficiency virus HIV-1. J Mol Biol. 1990 Dec 5;216(3):689–699. doi: 10.1016/0022-2836(90)90392-Y. [DOI] [PubMed] [Google Scholar]
  6. De Rocquigny H., Gabus C., Vincent A., Fournié-Zaluski M. C., Roques B., Darlix J. L. Viral RNA annealing activities of human immunodeficiency virus type 1 nucleocapsid protein require only peptide domains outside the zinc fingers. Proc Natl Acad Sci U S A. 1992 Jul 15;89(14):6472–6476. doi: 10.1073/pnas.89.14.6472. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Dorfman T., Luban J., Goff S. P., Haseltine W. A., Göttlinger H. G. Mapping of functionally important residues of a cysteine-histidine box in the human immunodeficiency virus type 1 nucleocapsid protein. J Virol. 1993 Oct;67(10):6159–6169. doi: 10.1128/jvi.67.10.6159-6169.1993. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Fields S., Song O. A novel genetic system to detect protein-protein interactions. Nature. 1989 Jul 20;340(6230):245–246. doi: 10.1038/340245a0. [DOI] [PubMed] [Google Scholar]
  9. Franke E. K., Yuan H. E., Bossolt K. L., Goff S. P., Luban J. Specificity and sequence requirements for interactions between various retroviral Gag proteins. J Virol. 1994 Aug;68(8):5300–5305. doi: 10.1128/jvi.68.8.5300-5305.1994. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Jacks T., Power M. D., Masiarz F. R., Luciw P. A., Barr P. J., Varmus H. E. Characterization of ribosomal frameshifting in HIV-1 gag-pol expression. Nature. 1988 Jan 21;331(6153):280–283. doi: 10.1038/331280a0. [DOI] [PubMed] [Google Scholar]
  11. James M. N., Sielecki A. R. Molecular structure of an aspartic proteinase zymogen, porcine pepsinogen, at 1.8 A resolution. Nature. 1986 Jan 2;319(6048):33–38. doi: 10.1038/319033a0. [DOI] [PubMed] [Google Scholar]
  12. Kaplan A. H., Manchester M., Swanstrom R. The activity of the protease of human immunodeficiency virus type 1 is initiated at the membrane of infected cells before the release of viral proteins and is required for release to occur with maximum efficiency. J Virol. 1994 Oct;68(10):6782–6786. doi: 10.1128/jvi.68.10.6782-6786.1994. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Kaplan A. H., Swanstrom R. Human immunodeficiency virus type 1 Gag proteins are processed in two cellular compartments. Proc Natl Acad Sci U S A. 1991 May 15;88(10):4528–4532. doi: 10.1073/pnas.88.10.4528. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. 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]
  15. Kräusslich H. G., Schneider H., Zybarth G., Carter C. A., Wimmer E. Processing of in vitro-synthesized gag precursor proteins of human immunodeficiency virus (HIV) type 1 by HIV proteinase generated in Escherichia coli. J Virol. 1988 Nov;62(11):4393–4397. doi: 10.1128/jvi.62.11.4393-4397.1988. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Laemmli U. K. Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature. 1970 Aug 15;227(5259):680–685. doi: 10.1038/227680a0. [DOI] [PubMed] [Google Scholar]
  17. Luban J., Goff S. P. Binding of human immunodeficiency virus type 1 (HIV-1) RNA to recombinant HIV-1 gag polyprotein. J Virol. 1991 Jun;65(6):3203–3212. doi: 10.1128/jvi.65.6.3203-3212.1991. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Luban J., Lee C., Goff S. P. Effect of linker insertion mutations in the human immunodeficiency virus type 1 gag gene on activation of viral protease expressed in bacteria. J Virol. 1993 Jun;67(6):3630–3634. doi: 10.1128/jvi.67.6.3630-3634.1993. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. Ma J., Ptashne M. A new class of yeast transcriptional activators. Cell. 1987 Oct 9;51(1):113–119. doi: 10.1016/0092-8674(87)90015-8. [DOI] [PubMed] [Google Scholar]
  20. Morellet N., Jullian N., De Rocquigny H., Maigret B., Darlix J. L., Roques B. P. Determination of the structure of the nucleocapsid protein NCp7 from the human immunodeficiency virus type 1 by 1H NMR. EMBO J. 1992 Aug;11(8):3059–3065. doi: 10.1002/j.1460-2075.1992.tb05377.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. Murray M. G., Hung W., Sadowski I., Das Mahapatra B. Inactivation of a yeast transactivator by the fused HIV-1 proteinase: a simple assay for inhibitors of the viral enzyme activity. Gene. 1993 Nov 30;134(1):123–128. doi: 10.1016/0378-1119(93)90185-6. [DOI] [PubMed] [Google Scholar]
  22. Partin K., Kräusslich H. G., Ehrlich L., Wimmer E., Carter C. Mutational analysis of a native substrate of the human immunodeficiency virus type 1 proteinase. J Virol. 1990 Aug;64(8):3938–3947. doi: 10.1128/jvi.64.8.3938-3947.1990. [DOI] [PMC free article] [PubMed] [Google Scholar]
  23. Partin K., Zybarth G., Ehrlich L., DeCrombrugghe M., Wimmer E., Carter C. Deletion of sequences upstream of the proteinase improves the proteolytic processing of human immunodeficiency virus type 1. Proc Natl Acad Sci U S A. 1991 Jun 1;88(11):4776–4780. doi: 10.1073/pnas.88.11.4776. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. Rhee S. S., Hunter E. Myristylation is required for intracellular transport but not for assembly of D-type retrovirus capsids. J Virol. 1987 Apr;61(4):1045–1053. doi: 10.1128/jvi.61.4.1045-1053.1987. [DOI] [PMC free article] [PubMed] [Google Scholar]
  25. Smith A. J., Cho M. I., Hammarskjöld M. L., Rekosh D. Human immunodeficiency virus type 1 Pr55gag and Pr160gag-pol expressed from a simian virus 40 late replacement vector are efficiently processed and assembled into viruslike particles. J Virol. 1990 Jun;64(6):2743–2750. doi: 10.1128/jvi.64.6.2743-2750.1990. [DOI] [PMC free article] [PubMed] [Google Scholar]
  26. South T. L., Blake P. R., Sowder R. C., 3rd, Arthur L. O., Henderson L. E., Summers M. F. The nucleocapsid protein isolated from HIV-1 particles binds zinc and forms retroviral-type zinc fingers. Biochemistry. 1990 Aug 28;29(34):7786–7789. doi: 10.1021/bi00486a002. [DOI] [PubMed] [Google Scholar]
  27. Studier F. W., Rosenberg A. H., Dunn J. J., Dubendorff J. W. Use of T7 RNA polymerase to direct expression of cloned genes. Methods Enzymol. 1990;185:60–89. doi: 10.1016/0076-6879(90)85008-c. [DOI] [PubMed] [Google Scholar]
  28. Van Hoy M., Leuther K. K., Kodadek T., Johnston S. A. The acidic activation domains of the GCN4 and GAL4 proteins are not alpha helical but form beta sheets. Cell. 1993 Feb 26;72(4):587–594. doi: 10.1016/0092-8674(93)90077-4. [DOI] [PubMed] [Google Scholar]
  29. Weber I. T. Comparison of the crystal structures and intersubunit interactions of human immunodeficiency and Rous sarcoma virus proteases. J Biol Chem. 1990 Jun 25;265(18):10492–10496. [PubMed] [Google Scholar]
  30. Wills J. W., Cameron C. E., Wilson C. B., Xiang Y., Bennett R. P., Leis J. An assembly domain of the Rous sarcoma virus Gag protein required late in budding. J Virol. 1994 Oct;68(10):6605–6618. doi: 10.1128/jvi.68.10.6605-6618.1994. [DOI] [PMC free article] [PubMed] [Google Scholar]
  31. Wills J. W., Craven R. C. Form, function, and use of retroviral gag proteins. AIDS. 1991 Jun;5(6):639–654. doi: 10.1097/00002030-199106000-00002. [DOI] [PubMed] [Google Scholar]
  32. 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]
  33. Wondrak E. M., Louis J. M., de Rocquigny H., Chermann J. C., Roques B. P. The gag precursor contains a specific HIV-1 protease cleavage site between the NC (P7) and P1 proteins. FEBS Lett. 1993 Oct 25;333(1-2):21–24. doi: 10.1016/0014-5793(93)80367-4. [DOI] [PubMed] [Google Scholar]
  34. Zybarth G., Kräusslich H. G., Partin K., Carter C. Proteolytic activity of novel human immunodeficiency virus type 1 proteinase proteins from a precursor with a blocking mutation at the N terminus of the PR domain. J Virol. 1994 Jan;68(1):240–250. doi: 10.1128/jvi.68.1.240-250.1994. [DOI] [PMC free article] [PubMed] [Google Scholar]