Giuseppe
Emeritus
[Source: Proceedings of the National Academy of Sciences of the United States of America, full page: (LINK). Abstract, edited.]
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Structure and inhibition of the drug-resistant S31N mutant of the M2 ion channel of influenza A virus
Jun Wang<SUP>a</SUP>, Yibing Wu<SUP>a</SUP>,<SUP>1</SUP>, Chunlong Ma<SUP>b</SUP>, Giacomo Fiorin<SUP>c</SUP>, Jizhou Wang<SUP>d</SUP>, Lawrence H. Pinto<SUP>e</SUP>, Robert A. Lamb<SUP>b</SUP>,<SUP>f</SUP>, Michael L. Klein<SUP>c</SUP>, and William F. DeGrado<SUP>a</SUP>,<SUP>1</SUP>
<SUP></SUP>
Author Affiliations: <SUP>a</SUP>Department of Pharmaceutical Chemistry, University of California, San Francisco, CA 94158-9001; Departments of <SUP>b</SUP>Molecular Biosciences and <SUP>e</SUP>Neurobiology, and <SUP>f</SUP>Howard Hughes Medical Institute, Northwestern University, Evanston, IL 60208-3500; <SUP>c</SUP>Institute for Computational Molecular Science and Department of Chemistry, Temple University, Philadelphia, PA 19122-6078; and <SUP>d</SUP>Influmedix, Inc., Radnor, PA 19087-5221
Edited by Adriaan Bax, National Institutes of Health, Bethesda, MD, and approved November 28, 2012 (received for review September 29, 2012)
Abstract
The influenza A virus M2 proton channel (A/M2) is the target of the antiviral drugs amantadine and rimantadine, whose use has been discontinued due to widespread drug resistance. Among the handful of drug-resistant mutants, S31N is found in more than 95% of the currently circulating viruses and shows greatly decreased inhibition by amantadine. The discovery of inhibitors of S31N has been hampered by the limited size, polarity, and dynamic nature of its amantadine-binding site. Nevertheless, we have discovered small-molecule drugs that inhibit S31N with potencies greater than amantadine?s potency against WT M2. Drug binding locks the protein into a well-defined conformation, and the NMR structure of the complex shows the drug bound in the homotetrameric channel, threaded between the side chains of Asn31. Unrestrained molecular dynamics simulations predicted the same binding site. This S31N inhibitor, like other potent M2 inhibitors, contains a charged ammonium group. The ammonium binds as a hydrate to one of three sites aligned along the central cavity that appear to be hotspots for inhibition. These sites might stabilize hydronium-like species formed as protons diffuse through the outer channel to the proton-shuttling residue His37 near the cytoplasmic end of the channel.
M2-S31N mutant structure - membrane protein structure - M2-S31N inhibitor
Footnotes
<SUP>1</SUP>To whom correspondence may be addressed. E-mail: yibing.wu@ucsf.edu or william.degrado@ucsf.edu.
Author contributions: Jun Wang, Y.W., and W.F.D. designed research; Jun Wang, Y.W., C.M., and G.F. performed research; Jun Wang, Y.W., C.M., G.F., Jizhou Wang, L.H.P., R.A.L., M.L.K., and W.F.D. contributed new reagents/analytic tools; Jun Wang, Y.W., and W.F.D. analyzed data; and Jun Wang, Y.W., and W.F.D. wrote the paper.
Conflict of interest statement: Jizhou Wang is an employee of Influmedix, Inc. W.F.D., R.A.L., M.L.K., and L.H.P. are members of the scientific advisory board of Influmedix, Inc.
This article is a PNAS Direct Submission.
Data deposition: The structure and chemical shift assignments have been deposited in the Protein Data Bank, www.pdb.org (accession no. 2LY0) and BioMagResBank, www.bmrb.wisc.edu (accession no. 18706), respectively.
This article contains supporting information online at www.pnas.org/lookup/suppl/doi:10.1073/pnas.1216526110/-/DCSupplemental.
-Jun Wang<SUP>a</SUP>, Yibing Wu<SUP>a</SUP>,<SUP>1</SUP>, Chunlong Ma<SUP>b</SUP>, Giacomo Fiorin<SUP>c</SUP>, Jizhou Wang<SUP>d</SUP>, Lawrence H. Pinto<SUP>e</SUP>, Robert A. Lamb<SUP>b</SUP>,<SUP>f</SUP>, Michael L. Klein<SUP>c</SUP>, and William F. DeGrado<SUP>a</SUP>,<SUP>1</SUP>
<SUP></SUP>
Author Affiliations: <SUP>a</SUP>Department of Pharmaceutical Chemistry, University of California, San Francisco, CA 94158-9001; Departments of <SUP>b</SUP>Molecular Biosciences and <SUP>e</SUP>Neurobiology, and <SUP>f</SUP>Howard Hughes Medical Institute, Northwestern University, Evanston, IL 60208-3500; <SUP>c</SUP>Institute for Computational Molecular Science and Department of Chemistry, Temple University, Philadelphia, PA 19122-6078; and <SUP>d</SUP>Influmedix, Inc., Radnor, PA 19087-5221
Edited by Adriaan Bax, National Institutes of Health, Bethesda, MD, and approved November 28, 2012 (received for review September 29, 2012)
Abstract
The influenza A virus M2 proton channel (A/M2) is the target of the antiviral drugs amantadine and rimantadine, whose use has been discontinued due to widespread drug resistance. Among the handful of drug-resistant mutants, S31N is found in more than 95% of the currently circulating viruses and shows greatly decreased inhibition by amantadine. The discovery of inhibitors of S31N has been hampered by the limited size, polarity, and dynamic nature of its amantadine-binding site. Nevertheless, we have discovered small-molecule drugs that inhibit S31N with potencies greater than amantadine?s potency against WT M2. Drug binding locks the protein into a well-defined conformation, and the NMR structure of the complex shows the drug bound in the homotetrameric channel, threaded between the side chains of Asn31. Unrestrained molecular dynamics simulations predicted the same binding site. This S31N inhibitor, like other potent M2 inhibitors, contains a charged ammonium group. The ammonium binds as a hydrate to one of three sites aligned along the central cavity that appear to be hotspots for inhibition. These sites might stabilize hydronium-like species formed as protons diffuse through the outer channel to the proton-shuttling residue His37 near the cytoplasmic end of the channel.
M2-S31N mutant structure - membrane protein structure - M2-S31N inhibitor
Footnotes
<SUP>1</SUP>To whom correspondence may be addressed. E-mail: yibing.wu@ucsf.edu or william.degrado@ucsf.edu.
Author contributions: Jun Wang, Y.W., and W.F.D. designed research; Jun Wang, Y.W., C.M., and G.F. performed research; Jun Wang, Y.W., C.M., G.F., Jizhou Wang, L.H.P., R.A.L., M.L.K., and W.F.D. contributed new reagents/analytic tools; Jun Wang, Y.W., and W.F.D. analyzed data; and Jun Wang, Y.W., and W.F.D. wrote the paper.
Conflict of interest statement: Jizhou Wang is an employee of Influmedix, Inc. W.F.D., R.A.L., M.L.K., and L.H.P. are members of the scientific advisory board of Influmedix, Inc.
This article is a PNAS Direct Submission.
Data deposition: The structure and chemical shift assignments have been deposited in the Protein Data Bank, www.pdb.org (accession no. 2LY0) and BioMagResBank, www.bmrb.wisc.edu (accession no. 18706), respectively.
This article contains supporting information online at www.pnas.org/lookup/suppl/doi:10.1073/pnas.1216526110/-/DCSupplemental.
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