Skip to main content
NCBI home page
Journal of Virology logoLink to Journal of Virology
. 1990 Aug;64(8):3824–3832. doi: 10.1128/jvi.64.8.3824-3832.1990

Conformational changes and fusion activity of influenza virus hemagglutinin of the H2 and H3 subtypes: effects of acid pretreatment.

A Puri 1, F P Booy 1, R W Doms 1, J M White 1, R Blumenthal 1
PMCID: PMC249678  PMID: 2196382

Abstract

Marked differences were observed between the H2 and H3 strains of influenza virus in their sensitivity to pretreatment at low pH. Whereas viral fusion and hemolysis mediated by influenza virus X:31 (H3 subtype) were inactivated by pretreatment of the virus at low pH, influenza virus A/Japan/305/57 (H2 subtype) retained those activities even after a 15-min incubation at pH 5.0 and 37 degrees C. Fusion with erythrocytes was measured by using the octadecylrhodamine-dequenching assay with both intact virions and CV-1 monkey kidney cells expressing hemagglutinin (HA) on the plasma membrane. To study the nature of the differences between the two strains, we examined the effects of low-pH treatment on the conformational change of HA by its susceptibility to protease digestion, exposure of the fusion peptide, and electron microscopy of unstained, frozen, hydrated virus. We found that the respective HA molecules from the two strains assumed different conformational states after exposure to low pH. The relationship between the conformation of HA and its fusogenic activity is discussed in the context of these experiments.

Full text

PDF
3824

Images in this article

3829Image
on p.3829
3830Image
on p.3830

Selected References

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

  1. Blumenthal R., Bali-Puri A., Walter A., Covell D., Eidelman O. pH-dependent fusion of vesicular stomatitis virus with Vero cells. Measurement by dequenching of octadecyl rhodamine fluorescence. J Biol Chem. 1987 Oct 5;262(28):13614–13619. [PubMed] [Google Scholar]
  2. Booy F. P., Ruigrok R. W., van Bruggen E. F. Electron microscopy of influenza virus. A comparison of negatively stained and ice-embedded particles. J Mol Biol. 1985 Aug 20;184(4):667–676. doi: 10.1016/0022-2836(85)90312-2. [DOI] [PubMed] [Google Scholar]
  3. Doms R. W., Helenius A., White J. Membrane fusion activity of the influenza virus hemagglutinin. The low pH-induced conformational change. J Biol Chem. 1985 Mar 10;260(5):2973–2981. [PubMed] [Google Scholar]
  4. Edwards J., Mann E., Brown D. T. Conformational changes in Sindbis virus envelope proteins accompanying exposure to low pH. J Virol. 1983 Mar;45(3):1090–1097. doi: 10.1128/jvi.45.3.1090-1097.1983. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Gething M. J., Bye J., Skehel J., Waterfield M. Cloning and DNA sequence of double-stranded copies of haemagglutinin genes from H2 and H3 strains elucidates antigenic shift and drift in human influenza virus. Nature. 1980 Sep 25;287(5780):301–306. doi: 10.1038/287301a0. [DOI] [PubMed] [Google Scholar]
  6. Gething M. J., Doms R. W., York D., White J. Studies on the mechanism of membrane fusion: site-specific mutagenesis of the hemagglutinin of influenza virus. J Cell Biol. 1986 Jan;102(1):11–23. doi: 10.1083/jcb.102.1.11. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Gollins S. W., Porterfield J. S. The uncoating and infectivity of the flavivirus West Nile on interaction with cells: effects of pH and ammonium chloride. J Gen Virol. 1986 Sep;67(Pt 9):1941–1950. doi: 10.1099/0022-1317-67-9-1941. [DOI] [PubMed] [Google Scholar]
  8. Hinshaw V. S., Webster R. G., Naeve C. W., Murphy B. R. Altered tissue tropism of human-avian reassortant influenza viruses. Virology. 1983 Jul 15;128(1):260–263. doi: 10.1016/0042-6822(83)90337-9. [DOI] [PubMed] [Google Scholar]
  9. Hoekstra D., Klappe K. Use of a fluorescence assay to monitor the kinetics of fusion between erythrocyte ghosts, as induced by Sendai virus. Biosci Rep. 1986 Nov;6(11):953–960. doi: 10.1007/BF01114971. [DOI] [PubMed] [Google Scholar]
  10. Hoekstra D., de Boer T., Klappe K., Wilschut J. Fluorescence method for measuring the kinetics of fusion between biological membranes. Biochemistry. 1984 Nov 20;23(24):5675–5681. doi: 10.1021/bi00319a002. [DOI] [PubMed] [Google Scholar]
  11. Junankar P. R., Cherry R. J. Temperature and pH dependence of the haemolytic activity of influenza virus and of the rotational mobility of the spike glycoproteins. Biochim Biophys Acta. 1986 Jan 29;854(2):198–206. doi: 10.1016/0005-2736(86)90111-2. [DOI] [PubMed] [Google Scholar]
  12. Klenk H. D., Rott R., Orlich M., Blödorn J. Activation of influenza A viruses by trypsin treatment. Virology. 1975 Dec;68(2):426–439. doi: 10.1016/0042-6822(75)90284-6. [DOI] [PubMed] [Google Scholar]
  13. Marsh M., Helenius A. Virus entry into animal cells. Adv Virus Res. 1989;36:107–151. doi: 10.1016/S0065-3527(08)60583-7. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. McClure M. O., Marsh M., Weiss R. A. Human immunodeficiency virus infection of CD4-bearing cells occurs by a pH-independent mechanism. EMBO J. 1988 Feb;7(2):513–518. doi: 10.1002/j.1460-2075.1988.tb02839.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Morris S. J., Sarkar D. P., White J. M., Blumenthal R. Kinetics of pH-dependent fusion between 3T3 fibroblasts expressing influenza hemagglutinin and red blood cells. Measurement by dequenching of fluorescence. J Biol Chem. 1989 Mar 5;264(7):3972–3978. [PubMed] [Google Scholar]
  16. OKADA Y. Analysis of giant polynuclear cell formation caused by HVJ virus from Ehrlich's ascites tumor cells. I. Microscopic observation of giant polynuclear cell formation. Exp Cell Res. 1962 Feb;26:98–107. doi: 10.1016/0014-4827(62)90205-7. [DOI] [PubMed] [Google Scholar]
  17. Puri A., Winick J., Lowy R. J., Covell D., Eidelman O., Walter A., Blumenthal R. Activation of vesicular stomatitis virus fusion with cells by pretreatment at low pH. J Biol Chem. 1988 Apr 5;263(10):4749–4753. [PubMed] [Google Scholar]
  18. Ruigrok R. W., Wrigley N. G., Calder L. J., Cusack S., Wharton S. A., Brown E. B., Skehel J. J. Electron microscopy of the low pH structure of influenza virus haemagglutinin. EMBO J. 1986 Jan;5(1):41–49. doi: 10.1002/j.1460-2075.1986.tb04175.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. Sarkar D. P., Morris S. J., Eidelman O., Zimmerberg J., Blumenthal R. Initial stages of influenza hemagglutinin-induced cell fusion monitored simultaneously by two fluorescent events: cytoplasmic continuity and lipid mixing. J Cell Biol. 1989 Jul;109(1):113–122. doi: 10.1083/jcb.109.1.113. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Sato S. B., Kawasaki K., Ohnishi S. Hemolytic activity of influenza virus hemagglutinin glycoproteins activated in mildly acidic environments. Proc Natl Acad Sci U S A. 1983 Jun;80(11):3153–3157. doi: 10.1073/pnas.80.11.3153. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. Sauter N. K., Bednarski M. D., Wurzburg B. A., Hanson J. E., Whitesides G. M., Skehel J. J., Wiley D. C. Hemagglutinins from two influenza virus variants bind to sialic acid derivatives with millimolar dissociation constants: a 500-MHz proton nuclear magnetic resonance study. Biochemistry. 1989 Oct 17;28(21):8388–8396. doi: 10.1021/bi00447a018. [DOI] [PubMed] [Google Scholar]
  22. Scholtissek C. Stability of infectious influenza A viruses at low pH and at elevated temperature. Vaccine. 1985 Sep;3(3 Suppl):215–218. doi: 10.1016/0264-410x(85)90109-4. [DOI] [PubMed] [Google Scholar]
  23. Skehel J. J., Bayley P. M., Brown E. B., Martin S. R., Waterfield M. D., White J. M., Wilson I. A., Wiley D. C. Changes in the conformation of influenza virus hemagglutinin at the pH optimum of virus-mediated membrane fusion. Proc Natl Acad Sci U S A. 1982 Feb;79(4):968–972. doi: 10.1073/pnas.79.4.968. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. Stegmann T., Booy F. P., Wilschut J. Effects of low pH on influenza virus. Activation and inactivation of the membrane fusion capacity of the hemagglutinin. J Biol Chem. 1987 Dec 25;262(36):17744–17749. [PubMed] [Google Scholar]
  25. White J. M. Viral and cellular membrane fusion proteins. Annu Rev Physiol. 1990;52:675–697. doi: 10.1146/annurev.ph.52.030190.003331. [DOI] [PubMed] [Google Scholar]
  26. White J. M., Wilson I. A. Anti-peptide antibodies detect steps in a protein conformational change: low-pH activation of the influenza virus hemagglutinin. J Cell Biol. 1987 Dec;105(6 Pt 2):2887–2896. doi: 10.1083/jcb.105.6.2887. [DOI] [PMC free article] [PubMed] [Google Scholar]
  27. White J., Kartenbeck J., Helenius A. Membrane fusion activity of influenza virus. EMBO J. 1982;1(2):217–222. doi: 10.1002/j.1460-2075.1982.tb01150.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  28. Wiley D. C., Skehel J. J. The structure and function of the hemagglutinin membrane glycoprotein of influenza virus. Annu Rev Biochem. 1987;56:365–394. doi: 10.1146/annurev.bi.56.070187.002053. [DOI] [PubMed] [Google Scholar]
  29. Yewdell J. W., Gerhard W., Bachi T. Monoclonal anti-hemagglutinin antibodies detect irreversible antigenic alterations that coincide with the acid activation of influenza virus A/PR/834-mediated hemolysis. J Virol. 1983 Oct;48(1):239–248. doi: 10.1128/jvi.48.1.239-248.1983. [DOI] [PMC free article] [PubMed] [Google Scholar]

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

RESOURCES