@article{0a114460d1bc4767840b9308a59e87f5,
title = "Proton transfer activity of the reconstituted Mycobacterium tuberculosis MmpL3 is modulated by substrate mimics and inhibitors",
abstract = "Transporters belonging to the Resistance-Nodulation-cell Division (RND) superfamily of proteins such as Mycobacterium tuberculosis MmpL3 and its analogs are the focus of intense investigations due to their importance in the physiology of Corynebacterium- Mycobacterium-Nocardia species and antimycobacterial drug discovery. These transporters deliver trehalose monomycolates, the precursors of major lipids of the outer membrane, to the periplasm by a proton motive force-dependent mechanism. In this study, we successfully purified, from native membranes, the full-length and the C-terminal truncated M. tuberculosis MmpL3 and Corynebacterium glutamicum CmpL1 proteins and reconstituted them into proteoliposomes. We also generated a series of substrate mimics and inhibitors specific to these transporters, analyzed their activities in the reconstituted proteoliposomes, and carried out molecular dynamics simulations of the model MmpL3 transporter at different pH. We found that all reconstituted proteins facilitate proton translocation across a phospholipid bilayer, but MmpL3 and CmpL1 differ dramatically in their responses to pH and interactions with substrate mimics and indole-2-carboxamide inhibitors. Our results further suggest that some inhibitors abolish the transport activity of MmpL3 and CmpL1 by inhibition of proton translocation.",
keywords = "Tuberculosis, drug target, membrane transporter, reconstitution",
author = "Stevens, {Casey M.} and Babii, {Svitlana O.} and Pandya, {Amitkumar N.} and Wei Li and Yupeng Li and Jitender Mehla and Robyn Scott and Pooja Hegde and Prathipati, {Pavan K.} and Atanu Acharya and Jinchan Liu and Gumbart, {James C.} and Jeffrey North and Mary Jackson and Zgurskaya, {Helen I.}",
note = "Funding Information: This work was supported by a grant from the NIH/National Institute of Allergy and Infectious Diseases (Grant AI116525) (to M.J., H.I.Z., J.N., and J.C.G). The content is solely the responsibility of the authors and does not necessarily represent the official views of the NIH. We are grateful to the Global Alliance for TB Drug Development for the provision of NITD-304 and NITD349, to Dr. Remuinan-Blanco (GSK Tres Cantos Open Lab Foundation) for the provision of THPP1, to Dr. Lee (St. Jude Children's Research Hospital) for the provision of SQ109, and to Dr. Jacobs (Albert Einstein College of Medicine, NY) for Mtb H37Rv strain mc26206. SPR experiments were carried out using a Biacore T200 instrument of the Oklahoma Medical Research Foundation Biacore Facility, which is funded by the Shared Instrumentation Grant S10 OD025014. Computational resources were provided through the Extreme Science and Engineering Discovery Environment (Grant TG-MCB130173), which is supported by NSF Grant ACI1548562. Y.L. and J.L. acknowledges Tang Aoqing Honors Program in Science, College of Chemistry, Jilin University, Changchun, Jilin Province, China. Funding Information: ACKNOWLEDGMENTS. This work was supported by a grant from the NIH/ National Institute of Allergy and Infectious Diseases (Grant AI116525) (to M.J., H.I.Z., J.N., and J.C.G). The content is solely the responsibility of the authors and does not necessarily represent the official views of the NIH. We are grateful to the Global Alliance for TB Drug Development for the provision of NITD-304 and NITD-349, to Dr. Remuinan-Blanco (GSK Tres Cantos Open Lab Foundation) for the provision of THPP1, to Dr. Lee (St. Jude Children{\textquoteright}s Research Hospital) for the provision of SQ109, and to Dr. Jacobs (Albert Einstein College of Medicine, NY) for Mtb H37Rv strain mc26206. SPR experiments were carried out using a Biacore T200 instrument of the Oklahoma Medical Research Foundation Biacore Facility, which is funded by the Shared Instrumentation Grant S10 OD025014. Computational resources were provided through the Extreme Science and Engineering Discovery Environment (Grant TG-MCB130173), which is supported by NSF Grant ACI-1548562. Y.L. and J.L. acknowledges Tang Aoqing Honors Program in Science, College of Chemistry, Jilin University, Changchun, Jilin Province, China. Publisher Copyright: Copyright {\textcopyright} 2022 the Author(s).",
year = "2022",
month = jul,
day = "26",
doi = "10.1073/pnas.2113963119",
language = "English (US)",
volume = "119",
journal = "Proceedings of the National Academy of Sciences of the United States of America",
issn = "0027-8424",
publisher = "National Academy of Sciences",
number = "30",
}