title: Kinetics of proton transport into influenza virions by the viral M2 channel.
abstract: M2 protein of influenza A viruses is a tetrameric transmembrane proton channel, which has essential functions both early and late in the virus infectious cycle. Previous studies of proton transport by M2 have been limited to measurements outside the context of the virus particle. We have developed an in vitro fluorescence-based assay to monitor internal acidification of individual virions triggered to undergo membrane fusion. We show that rimantadine, an inhibitor of M2 proton conductance, blocks the acidification-dependent dissipation of fluorescence from a pH-sensitive virus-content probe. Fusion-pore formation usually follows internal acidification but does not require it. The rate of internal virion acidification increases with external proton concentration and saturates with a pK(m) of ∼4.7. The rate of proton transport through a single, fully protonated M2 channel is approximately 100 to 400 protons per second. The saturating proton-concentration dependence and the low rate of internal virion acidification derived from authentic virions support a transporter model for the mechanism of proton transfer.
authors.affiliation: Department of Biological Chemistry and Molecular Pharmacology, Harvard Medical School, Boston, Massachusetts, United States of America.
authors.forename: Tijana
authors.initials: T
authors.lastname: Ivanovic
authors.forename: Rutger
authors.initials: R
authors.lastname: Rozendaal
authors.forename: Daniel L
authors.initials: DL
authors.lastname: Floyd
authors.forename: Milos
authors.initials: M
authors.lastname: Popovic
authors.forename: Antoine M
authors.initials: AM
authors.lastname: van Oijen
authors.forename: Stephen C
authors.initials: SC
authors.lastname: Harrison
mesh_terms: D000998:Antiviral Agents / Q000494:pharmacology; D019793:Fluorescein / Q000378:metabolism; D006863:Hydrogen-Ion Concentration; D009980:Influenza A virus / Q000187:drug effects / Q000378:metabolism*; D017136:Ion Transport / Q000187:drug effects / Q000502:physiology*; D007700:Kinetics; D011522:Protons; D012299:Rimantadine / Q000494:pharmacology; D014763:Viral Matrix Proteins / Q000378:metabolism*; D014771:Virion / Q000187:drug effects / Q000378:metabolism*; D053586:Virus Internalization / Q000187:drug effects; D000086082:Viroporin Proteins
chemical_list: D000998:Antiviral Agents; D019793:Fluorescein; D011522:Protons; D012299:Rimantadine; D014763:Viral Matrix Proteins; C488937:M2 protein, Influenza A virus; D000086082:Viroporin Proteins