Kinetics and pH dependence of acid-induced structural changes in the lymphocytic choriomeningitis virus glycoprotein complex.
The NBD-PE/Rd-PE fluorescent membrane fusion assay was used to measure the pH dependence and kinetics of the fusion activity of lymphocytic choriomeningitis virus with liposomes designed to mimic the composition of the endosomal membrane. Fusion activity was only observed at pH values less than 6.3 and showed a greater rate and extent at lower pH values. Pronounced kinetic fusion curves were observed at pH values below 5.8. When equivalent lipid amounts of target liposomes and virus were mixed at pH 5.3 the dequenching activity had a t1/2 of 45 +/- 10 sec. In addition to catalyzing membrane fusion after acidification the glycoprotein complex was previously found to undergo conformational change (C. Di Simone, M. A. Zandonatti, and M. J. Buchmeier, 1994, Virology 198, 455-465), including loss of the GP-1 polypeptide from the virion surface. The pH dependence and kinetics of this acid-induced GP-1 release were quantitated using centrifugal separation of solubilized GP-1 from pelleted virions. A pH-dependent elution curve was determined with progressively more GP-1 released at pH values below 6.3 and reaching nearly 100% dissociation at pH 5.5 after 30 min at 37 degrees. At pH 5.3 the GP disassembly proceeded with a t1/2 of 7 +/- 2 min. The t1/2 of virus inactivation was also measured at pH 5.3 and 7.0 and found to be 7.9 +/- 1 and 150 min, respectively. Fusion, GP dissociation, and inactivation kinetics data suggest a mechanism in which GP is activated to a fusion active state where membrane lipid exchange occurs and then undergoes an irreversible conformational change which includes the loss of GP-1 from the spike complex.
Di Simone
C
C
Department of Neuropharmacology, Scripps Research Institute, La Jolla, California 92037, USA.
Buchmeier
M J
MJ
D000818:Animals; D006801:Humans; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D008081:Liposomes; D008216:Lymphocytic Choriomeningitis / Q000209:etiology; D008217:Lymphocytic choriomeningitis virus / Q000737:chemistry* / Q000235:genetics / Q000472:pathogenicity; D008561:Membrane Fusion; D008562:Membrane Glycoproteins / Q000737:chemistry*; D015394:Molecular Structure; D014764:Viral Proteins / Q000737:chemistry*
D008081:Liposomes; D008562:Membrane Glycoproteins; D014764:Viral Proteins