An amantadine-sensitive chimeric BM2 ion channel of influenza B virus has implications for the mechanism of drug inhibition.
Influenza A virus M2 (A/M2) and the influenza B virus BM2 are both small integral membrane proteins that form proton-selective ion channels. Influenza A virus A/M2 channel is the target of the antiviral drug amantadine (and its methyl derivative rimantadine), whereas BM2 channel activity is not affected by the drug. The atomic structure of the pore-transmembrane (TM) domain peptide has been determined by x-ray crystallography [Stouffer et al. (2008) Nature 451:596-599] and of a larger M2 peptide by NMR methods [Schnell and Chou (2008) Nature 451:591-595]. The crystallographic data show electron density (at 3.5 A resolution) in the channel pore, consistent with amantadine blocking the pore of the channel. In contrast, the NMR data show 4 rimantadine molecules bound on the outside of the helices toward the cytoplasmic side of the membrane. Drug binding includes interactions with residues 40-45 and a polar hydrogen bond between rimantadine and aspartic acid residue 44 (D44). These 2 distinct drug-binding sites led to 2 incompatible drug inhibition mechanisms. We have generated chimeric channels between amantadine-sensitive A/M2 and amantadine-insensitive BM2 designed to define the drug-binding site. Two chimeras containing 5 residues of the A/M2 ectodomain and residues 24-36 of the A/M2 TM domain show 85% amantadine/rimantadine sensitivity and specific activity comparable to that of WT BM2. These functional data suggest that the amantadine/rimantadine binding site identified on the outside of the 4 helices is not the primary site associated with the pharmacologic inhibition of the A/M2 ion channel.
Ohigashi
Yuki
Y
Department of Neurobiology and Physiology, Northwestern University, Evanston, IL 60208-3500, USA.
Ma
Chunlong
C
Jing
Xianghong
X
Balannick
Victoria
V
Pinto
Lawrence H
LH
Lamb
Robert A
RA
D000547:Amantadine / Q000494:pharmacology*; D000595:Amino Acid Sequence; D019943:Amino Acid Substitution / Q000187:drug effects; D004305:Dose-Response Relationship, Drug; D006863:Hydrogen-Ion Concentration / Q000187:drug effects; D009981:Influenza B virus / Q000187:drug effects*; D015640:Ion Channel Gating / Q000187:drug effects; D007473:Ion Channels / Q000037:antagonists & inhibitors* / Q000737:chemistry / Q000378:metabolism; D008969:Molecular Sequence Data; D009154:Mutation / Q000235:genetics; D011994:Recombinant Proteins / Q000737:chemistry / Q000378:metabolism*; D014764:Viral Proteins / Q000037:antagonists & inhibitors* / Q000737:chemistry / Q000378:metabolism
C064964:BM2 polypeptide, influenza B virus; D007473:Ion Channels; D011994:Recombinant Proteins; D014764:Viral Proteins; D000547:Amantadine