@article{227a9706fee5422a9711ac7f9493cd8c,
title = "Critical Role of Water Molecules in Proton Translocation by the Membrane-Bound Transhydrogenase",
abstract = "The nicotinamide nucleotide transhydrogenase (TH) is an integral membrane enzyme that uses the proton-motive force to drive hydride transfer from NADH to NADP+ in bacteria and eukaryotes. Here we solved a 2.2-{\AA} crystal structure of the TH transmembrane domain (Thermus thermophilus) at pH 6.5. This structure exhibits conformational changes of helix positions from a previous structure solved at pH 8.5, and reveals internal water molecules interacting with residues implicated in proton translocation. Together with molecular dynamics simulations, we show that transient water flows across a narrow pore and a hydrophobic “dry” region in the middle of the membrane channel, with key residues His42α2 (chain A) being protonated and Thr214β (chain B) displaying a conformational change, respectively, to gate the channel access to both cytoplasmic and periplasmic chambers. Mutation of Thr214β to Ala deactivated the enzyme. These data provide new insights into the gating mechanism of proton translocation in TH.",
keywords = "X-ray crystallography, membrane protein, molecular dynamics simulations, proton channel, transhydrogenase",
author = "Padayatti, \{Pius S.\} and Leung, \{Josephine H.\} and Paween Mahinthichaichan and Emad Tajkhorshid and Andrii Ishchenko and Vadim Cherezov and Soltis, \{S. Michael\} and Jackson, \{J. Baz\} and Stout, \{C. David\} and Gennis, \{Robert B.\} and Qinghai Zhang",
note = "We would like to dedicate this paper to the loving memory of Dr. C. David Stout (1947–2016) who enormously influenced all of us and was the driving force behind the understanding of transhydrogenase membrane protein structural studies. We thank synchrotron staff Sheila Trznadel, Craig Ogata, and Steven Corcoran at APS (23ID-D and 24ID-C); David Neau of NECAT, Lisa Dunn, Doukov Tzanko and Mathews Irimpan of SSRL at experimental station 12-2 for generous beamtime help; and Eric Johnson and Duncan McRee for comments on the manuscript. This work is supported by NIH grants 5R01GM103838 (to Q.Z.) and R01HL16101 (to R.B.G.). The atomic coordinates and structure factors have been deposited in the RCSB PDB (PDB: 5UNI). MD simulation resources were provided to R.B.G. by the startup XSEDE grant TG-MCB150029. Use of Advanced Photon Source at GM/CA@APS has been funded in whole or in part with Federal funds from the National Cancer Institute (ACB-12002) and the National Institute of General Medical Sciences (AGM-12006). This research used resources of the Advanced Photon Source, a U.S. Department of Energy (DOE) Office of Science User Facility operated for the DOE Office of Science by Argonne National Laboratory under contract no. DE-AC02-06CH11357. The Eiger 16M detector was funded by an NIH—Office of Research Infrastructure Programs, High-End Instrumentation grant (1S10OD012289-01A1). The SSRL Structural Molecular Biology Program is supported by the DOE Office of Biological and Environmental Research and by the NIGMS (including grant P41GM103393).",
year = "2017",
month = jul,
day = "5",
doi = "10.1016/j.str.2017.05.022",
language = "English (US)",
volume = "25",
pages = "1111--1119.e3",
journal = "Structure",
issn = "0969-2126",
publisher = "Cell Press",
number = "7",
}