XFEL structures of the influenza M2 proton channel: Room temperature water networks and insights into proton conduction

Jessica L. Thomaston, Rahel A. Woldeyes, Takanori Nakane, Ayumi Yamashita, Tomoyuki Tanaka, Kotaro Koiwai, Aaron S. Brewster, Benjamin A. Barad, Yujie Chen, Thomas Lemmin, Monarin Uervirojnangkoorn, Toshi Arima, Jun Kobayashi, Tetsuya Masuda, Mamoru Suzuki, Michihiro Sugahara, Nicholas K. Sauter, Rie Tanaka, Osamu Nureki, Kensuke TonoYasumasa Joti, Eriko Nango, So Iwata, Fumiaki Yumoto, James S. Fraser, William F. DeGrado

Research output: Contribution to journalArticlepeer-review

57 Citations (Scopus)


TheM2 proton channel of influenza A is a drug target that is essential for the reproduction of the flu virus. It is also a model system for the study of selective, unidirectional proton transport across amembrane. Ordered water molecules arranged in "wires" inside the channel pore have been proposed to play a role in both the conduction of protons to the four gating His37 residues and the stabilization of multiple positive charges within the channel. To visualize the solvent in the pore of the channel at room temperature while minimizing the effects of radiation damage, data were collected to a resolution of 1.4 Å using an X-ray free-electron laser (XFEL) at three different pH conditions: pH 5.5, pH 6.5, and pH 8.0. Data were collected on the Inwardopen state, which is an intermediate that accumulates at high protonation of the His37 tetrad. At pH 5.5, a continuous hydrogen-bonded network of water molecules spans the vertical length of the channel, consistent with a Grotthuss mechanism model for proton transport to the His37 tetrad. This ordered solvent at pH 5.5 could act to stabilize the positive charges that build up on the gating His37 tetrad during the proton conduction cycle. The number of ordered pore waters decreases at pH 6.5 and 8.0, where the Inwardopen state is less stable. These studies provide a graphical view of the response of water to a change in charge within a restricted channel environment.

Original languageEnglish
Pages (from-to)13357-13362
Number of pages6
JournalProceedings of the National Academy of Sciences of the United States of America
Issue number51
Publication statusPublished - 2017 Dec 19


  • Influenza
  • Membrane protein
  • Proton channel
  • XFEL


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