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Structure of the amantadine binding site of influenza M2 proton channels in lipid bilayers

The M2 protein of influenza A virus is a membrane-spanning tetrameric proton channel targeted by the antiviral drugs amantadine and rimantadine (1). Resistance to these drugs has compromised their effectiveness against many influenza strains, including pandemic H1N1. A recent crystal structure of M2... Full description

Journal Title: Nature 2010, Vol.463(7281), p.689
Main Author: Sarah D. Cady
Other Authors: Klaus Schmidt-Rohr , Jun Wang , Cinque S. Soto , William F. Degrado , Mei Hong
Format: Electronic Article Electronic Article
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ID: ISSN: 0028-0836 ; E-ISSN: 1476-4687 ; DOI: 10.1038/nature08722
Link: http://dx.doi.org/10.1038/nature08722
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recordid: nature_a10.1038/nature08722
title: Structure of the amantadine binding site of influenza M2 proton channels in lipid bilayers
format: Article
creator:
  • Sarah D. Cady
  • Klaus Schmidt-Rohr
  • Jun Wang
  • Cinque S. Soto
  • William F. Degrado
  • Mei Hong
subjects:
  • Crystal Structure -- Analysis
  • Nuclear Magnetic Resonance Spectroscopy -- Usage
  • Proteins -- Structure
  • Membrane Proteins -- Chemical Properties
  • Antiviral Agents -- Chemical Properties
ispartof: Nature, 2010, Vol.463(7281), p.689
description: The M2 protein of influenza A virus is a membrane-spanning tetrameric proton channel targeted by the antiviral drugs amantadine and rimantadine (1). Resistance to these drugs has compromised their effectiveness against many influenza strains, including pandemic H1N1. A recent crystal structure of M2(22-46) showed electron densities attributed to a single amantadine in the amino-terminal half of the pore (2), indicating a physical occlusion mechanism for inhibition. However, a solution NMR structure of M2(18-60) showed four rimantadines bound to the carboxy-terminal lipid-facing surface of the helices (3), suggesting an allosteric mechanism. Here we show by solid-state NMR spectroscopy that two amantadine-binding sites exist in M2 in phospholipid bilayers. The high-affinity site, occupied by a single amantadine, is located in the N-terminal channel lumen, surrounded by residues mutated in amantadine-resistant viruses. Quantification of the protein-amantadine distances resulted in a 0.3 [Angstrom]-resolution structure of the high-affinity binding site. The second, low-affinity, site was observed on the C-terminal protein surface, but only when the drug reaches high concentrations in the bilayer. The orientation and dynamics of the drug are distinct in the two sites, as shown by [sup.2]H NMR. These results indicate that amantadine physically occludes the M2 channel, thus paving the way for developing new antiviral drugs against influenza viruses. The study demonstrates the ability of solid-state NMR to elucidate small-molecule interactions with membrane proteins and determine high-resolution structures of their complexes.
language:
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identifier: ISSN: 0028-0836 ; E-ISSN: 1476-4687 ; DOI: 10.1038/nature08722
fulltext: fulltext
issn:
  • 0028-0836
  • 00280836
  • 1476-4687
  • 14764687
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titleStructure of the amantadine binding site of influenza M2 proton channels in lipid bilayers
creatorSarah D. Cady ; Klaus Schmidt-Rohr ; Jun Wang ; Cinque S. Soto ; William F. Degrado ; Mei Hong
ispartofNature, 2010, Vol.463(7281), p.689
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descriptionThe M2 protein of influenza A virus is a membrane-spanning tetrameric proton channel targeted by the antiviral drugs amantadine and rimantadine (1). Resistance to these drugs has compromised their effectiveness against many influenza strains, including pandemic H1N1. A recent crystal structure of M2(22-46) showed electron densities attributed to a single amantadine in the amino-terminal half of the pore (2), indicating a physical occlusion mechanism for inhibition. However, a solution NMR structure of M2(18-60) showed four rimantadines bound to the carboxy-terminal lipid-facing surface of the helices (3), suggesting an allosteric mechanism. Here we show by solid-state NMR spectroscopy that two amantadine-binding sites exist in M2 in phospholipid bilayers. The high-affinity site, occupied by a single amantadine, is located in the N-terminal channel lumen, surrounded by residues mutated in amantadine-resistant viruses. Quantification of the protein-amantadine distances resulted in a 0.3 [Angstrom]-resolution structure of the high-affinity binding site. The second, low-affinity, site was observed on the C-terminal protein surface, but only when the drug reaches high concentrations in the bilayer. The orientation and dynamics of the drug are distinct in the two sites, as shown by [sup.2]H NMR. These results indicate that amantadine physically occludes the M2 channel, thus paving the way for developing new antiviral drugs against influenza viruses. The study demonstrates the ability of solid-state NMR to elucidate small-molecule interactions with membrane proteins and determine high-resolution structures of their complexes.
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