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. 1997 Feb;71(2):1107–1114. doi: 10.1128/jvi.71.2.1107-1114.1997

Topology of the large envelope protein of duck hepatitis B virus suggests a mechanism for membrane translocation during particle morphogenesis.

J T Guo 1, J C Pugh 1
PMCID: PMC191162  PMID: 8995631

Abstract

We have investigated the membrane topology of the large envelope protein of duck hepatitis B virus (DHBV) by protease protection and Western blot analysis, using monoclonal antibodies specific for the pre-S and S regions of the DHBV envelope to characterize protease-resistant polypeptides. These studies showed that DHBV L protein exhibits a mixed membrane topology similar to that of human hepatitis B virus L, with approximately half of the L molecules displaying pre-S on the surface of virus particles and the remainder with pre-S sequestered inside the virus envelope. The C-terminal region of DHBV pre-S was susceptible to protease digestion on all DHBV particle L protein, indicating that this region was externally disposed. DHBV L protein pre-S was entirely cytosolic immediately after synthesis. Our data, therefore, suggested that an intermediate form of the DHBV L molecule exists in mature envelope particles in which L is partially translocated or exists in a translocation-ready conformation. Incubation of virus particles at low pH and 37 degrees C triggered conversion of this intermediate into a fully translocated form. We have proposed a model for pre-S translocation based on our results that invokes the presence of an aqueous pore in the virus envelope, most likely created by oligomerization of transmembrane domains in the S region. The model predicts that pre-S is transported through this pore and that a loop structure is formed because the N terminus remains anchored to the inner face of the membrane. This translocation process occurs during particle morphogenesis and may also be a prerequisite to virus uncoating during infection.

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Selected References

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  1. Bruss V., Lu X., Thomssen R., Gerlich W. H. Post-translational alterations in transmembrane topology of the hepatitis B virus large envelope protein. EMBO J. 1994 May 15;13(10):2273–2279. doi: 10.1002/j.1460-2075.1994.tb06509.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Bruss V., Thomssen R. Mapping a region of the large envelope protein required for hepatitis B virion maturation. J Virol. 1994 Mar;68(3):1643–1650. doi: 10.1128/jvi.68.3.1643-1650.1994. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Bruss V., Vieluf K. Functions of the internal pre-S domain of the large surface protein in hepatitis B virus particle morphogenesis. J Virol. 1995 Nov;69(11):6652–6657. doi: 10.1128/jvi.69.11.6652-6657.1995. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Condreay L. D., Aldrich C. E., Coates L., Mason W. S., Wu T. T. Efficient duck hepatitis B virus production by an avian liver tumor cell line. J Virol. 1990 Jul;64(7):3249–3258. doi: 10.1128/jvi.64.7.3249-3258.1990. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Driessen A. J. Bacterial protein translocation: kinetic and thermodynamic role of ATP and the protonmotive force. Trends Biochem Sci. 1992 Jun;17(6):219–223. doi: 10.1016/0968-0004(92)90381-i. [DOI] [PubMed] [Google Scholar]
  6. Eble B. E., Lingappa V. R., Ganem D. Hepatitis B surface antigen: an unusual secreted protein initially synthesized as a transmembrane polypeptide. Mol Cell Biol. 1986 May;6(5):1454–1463. doi: 10.1128/mcb.6.5.1454. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Gilbert J. M., Hernandez L. D., Balliet J. W., Bates P., White J. M. Receptor-induced conformational changes in the subgroup A avian leukosis and sarcoma virus envelope glycoprotein. J Virol. 1995 Dec;69(12):7410–7415. doi: 10.1128/jvi.69.12.7410-7415.1995. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Gilmore R. Protein translocation across the endoplasmic reticulum: a tunnel with toll booths at entry and exit. Cell. 1993 Nov 19;75(4):589–592. doi: 10.1016/0092-8674(93)90476-7. [DOI] [PubMed] [Google Scholar]
  9. Helenius A. Unpacking the incoming influenza virus. Cell. 1992 May 15;69(4):577–578. doi: 10.1016/0092-8674(92)90219-3. [DOI] [PubMed] [Google Scholar]
  10. Klingmüller U., Schaller H. Hepadnavirus infection requires interaction between the viral pre-S domain and a specific hepatocellular receptor. J Virol. 1993 Dec;67(12):7414–7422. doi: 10.1128/jvi.67.12.7414-7422.1993. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Kuhn A., Zhu H. Y., Dalbey R. E. Efficient translocation of positively charged residues of M13 procoat protein across the membrane excludes electrophoresis as the primary force for membrane insertion. EMBO J. 1990 Aug;9(8):2385–2389. doi: 10.1002/j.1460-2075.1990.tb07413.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Lenhoff R. J., Summers J. Coordinate regulation of replication and virus assembly by the large envelope protein of an avian hepadnavirus. J Virol. 1994 Jul;68(7):4565–4571. doi: 10.1128/jvi.68.7.4565-4571.1994. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Mandart E., Kay A., Galibert F. Nucleotide sequence of a cloned duck hepatitis B virus genome: comparison with woodchuck and human hepatitis B virus sequences. J Virol. 1984 Mar;49(3):782–792. doi: 10.1128/jvi.49.3.782-792.1984. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Martoglio B., Dobberstein B. Snapshots of membrane-translocating proteins. Trends Cell Biol. 1996 Apr;6(4):142–147. doi: 10.1016/0962-8924(96)10001-5. [DOI] [PubMed] [Google Scholar]
  15. Neurath A. R., Kent S. B., Strick N., Parker K. Identification and chemical synthesis of a host cell receptor binding site on hepatitis B virus. Cell. 1986 Aug 1;46(3):429–436. doi: 10.1016/0092-8674(86)90663-x. [DOI] [PubMed] [Google Scholar]
  16. Offensperger W. B., Offensperger S., Walter E., Blum H. E., Gerok W. Inhibition of duck hepatitis B virus infection by lysosomotropic agents. Virology. 1991 Jul;183(1):415–418. doi: 10.1016/0042-6822(91)90157-7. [DOI] [PubMed] [Google Scholar]
  17. Ostapchuk P., Hearing P., Ganem D. A dramatic shift in the transmembrane topology of a viral envelope glycoprotein accompanies hepatitis B viral morphogenesis. EMBO J. 1994 Mar 1;13(5):1048–1057. doi: 10.1002/j.1460-2075.1994.tb06353.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Persing D. H., Varmus H. E., Ganem D. The preS1 protein of hepatitis B virus is acylated at its amino terminus with myristic acid. J Virol. 1987 May;61(5):1672–1677. doi: 10.1128/jvi.61.5.1672-1677.1987. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. Pontisso P., Petit M. A., Bankowski M. J., Peeples M. E. Human liver plasma membranes contain receptors for the hepatitis B virus pre-S1 region and, via polymerized human serum albumin, for the pre-S2 region. J Virol. 1989 May;63(5):1981–1988. doi: 10.1128/jvi.63.5.1981-1988.1989. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Prange R., Streeck R. E. Novel transmembrane topology of the hepatitis B virus envelope proteins. EMBO J. 1995 Jan 16;14(2):247–256. doi: 10.1002/j.1460-2075.1995.tb06998.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. Pugh J. C., Di Q., Mason W. S., Simmons H. Susceptibility to duck hepatitis B virus infection is associated with the presence of cell surface receptor sites that efficiently bind viral particles. J Virol. 1995 Aug;69(8):4814–4822. doi: 10.1128/jvi.69.8.4814-4822.1995. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Pugh J. C., Sninsky J. J., Summers J. W., Schaeffer E. Characterization of a pre-S polypeptide on the surfaces of infectious avian hepadnavirus particles. J Virol. 1987 May;61(5):1384–1390. doi: 10.1128/jvi.61.5.1384-1390.1987. [DOI] [PMC free article] [PubMed] [Google Scholar]
  23. Rigg R. J., Schaller H. Duck hepatitis B virus infection of hepatocytes is not dependent on low pH. J Virol. 1992 May;66(5):2829–2836. doi: 10.1128/jvi.66.5.2829-2836.1992. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. Schlicht H. J., Kuhn C., Guhr B., Mattaliano R. J., Schaller H. Biochemical and immunological characterization of the duck hepatitis B virus envelope proteins. J Virol. 1987 Jul;61(7):2280–2285. doi: 10.1128/jvi.61.7.2280-2285.1987. [DOI] [PMC free article] [PubMed] [Google Scholar]
  25. Simon K., Lingappa V. R., Ganem D. Secreted hepatitis B surface antigen polypeptides are derived from a transmembrane precursor. J Cell Biol. 1988 Dec;107(6 Pt 1):2163–2168. doi: 10.1083/jcb.107.6.2163. [DOI] [PMC free article] [PubMed] [Google Scholar]
  26. Stirk H. J., Thornton J. M., Howard C. R. A topological model for hepatitis B surface antigen. Intervirology. 1992;33(3):148–158. doi: 10.1159/000150244. [DOI] [PubMed] [Google Scholar]
  27. Summers J., Smith P. M., Huang M. J., Yu M. S. Morphogenetic and regulatory effects of mutations in the envelope proteins of an avian hepadnavirus. J Virol. 1991 Mar;65(3):1310–1317. doi: 10.1128/jvi.65.3.1310-1317.1991. [DOI] [PMC free article] [PubMed] [Google Scholar]
  28. Tuttleman J. S., Pugh J. C., Summers J. W. In vitro experimental infection of primary duck hepatocyte cultures with duck hepatitis B virus. J Virol. 1986 Apr;58(1):17–25. doi: 10.1128/jvi.58.1.17-25.1986. [DOI] [PMC free article] [PubMed] [Google Scholar]
  29. Yuasa S., Cheung R. C., Pham Q., Robinson W. S., Marion P. L. Peptide mapping of neutralizing and nonneutralizing epitopes of duck hepatitis B virus pre-S polypeptide. Virology. 1991 Mar;181(1):14–21. doi: 10.1016/0042-6822(91)90465-n. [DOI] [PubMed] [Google Scholar]

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