The oligomeric state of the active BM2 ion channel protein of influenza B virus.
Influenza A virus and influenza B virus particles both contain small integral membrane proteins (A/M2 and BM2, respectively) that function as a pH-sensitive proton channel and are essential for virus replication. The mechanism of action of the M2 channels is a subject of scientific interest particularly as A/M2 channel was shown to be a target for the action of the antiviral drug amantadine. Unfortunately, an inhibitor of the BM2 channel activity is not known. Thus, knowledge of the structural and functional properties of the BM2 channel is essential for the development of potent antiviral drugs. The characterization of the oligomeric state of the BM2 channel is an essential first step in the understanding of channel function. Here we describe determination of the stoichiometry of the BM2 proton channel by utilizing three different approaches. 1) We demonstrated that BM2 monomers can be chemically cross-linked to yield species consistent with dimers, trimers, and tetramers. 2) We studied electrophysiological and biochemical properties of mixed oligomers consisting of wild-type and mutated BM2 subunits and related these data to predicted binomial distribution models. 3) We used fluorescence resonance energy transfer (FRET) in combination with biochemical measurements to estimate the relationships between BM2 channel subunits expressed in the plasma membrane. Our experimental data are consistent with a tetrameric structure of the BM2 channel. Finally, we demonstrated that BM2 transmembrane domain is responsible for the channel oligomerization.
Balannik
Victoria
V
Department of Neurobiology and Physiology, Howard Hughes Medical Institute, Northwestern University, Evanston, IL 60208-3500, USA.
Lamb
Robert A
RA
Pinto
Lawrence H
LH
D000547:Amantadine / Q000494:pharmacology; D000998:Antiviral Agents / Q000494:pharmacology; D002460:Cell Line; D002462:Cell Membrane / Q000235:genetics / Q000378:metabolism*; D031541:Fluorescence Resonance Energy Transfer; D006801:Humans; D009980:Influenza A virus / Q000235:genetics / Q000378:metabolism; D009981:Influenza B virus / Q000502:physiology*; D007473:Ion Channels / Q000235:genetics / Q000378:metabolism*; D009154:Mutation; D020836:Protein Structure, Quaternary; D017434:Protein Structure, Tertiary; D021122:Protein Subunits; D013329:Structure-Activity Relationship; D014763:Viral Matrix Proteins / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism; D014764:Viral Proteins / Q000235:genetics / Q000378:metabolism*; D014779:Virus Replication / Q000187:drug effects / Q000502:physiology*; D000086082:Viroporin Proteins
D000547:Amantadine; D000998:Antiviral Agents; D007473:Ion Channels; D021122:Protein Subunits; D014763:Viral Matrix Proteins; D014764:Viral Proteins; C064964:BM2 polypeptide, influenza B virus; C488937:M2 protein, Influenza A virus; D000086082:Viroporin Proteins