Jin, Y., Molt Jr., R.W., Pellegrini, E. et al. (5 more authors) (2017) Assessing the influence of mutation on GTPase transition states using X-ray, 19F NMR, and DFT approaches. Angewandte Chemie International Edition, 56 (33). pp. 9732-9735. ISSN 1433-7851
Abstract
We report X-ray crystallographic and 19F NMR studies of the G-protein RhoA complexed with MgF3−, GDP, and RhoGAP, which has the mutation Arg85′Ala. When combined with DFT calculations, these data permit the identification of changes in transition state (TS) properties. The X-ray data show how Tyr34 maintains solvent exclusion and the core H-bond network in the active site by relocating to replace the missing Arg85′ sidechain. The 19F NMR data show deshielding effects that indicate the main function of Arg85′ is electronic polarization of the transferring phosphoryl group, primarily mediated by H-bonding to O3G and thence to PG. DFT calculations identify electron-density redistribution and pinpoint why the TS for guanosine 5′-triphosphate (GTP) hydrolysis is higher in energy when RhoA is complexed with RhoGAPArg85′Ala relative to wild-type (WT) RhoGAP. This study demonstrates that 19F NMR measurements, in combination with X-ray crystallography and DFT calculations, can reliably dissect the response of small GTPases to site-specific modification
Metadata
Item Type: | Article |
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Authors/Creators: |
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Copyright, Publisher and Additional Information: | © 2017 The Authors. Published by Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim. This is an open access article under the terms of the Creative Commons Attribution License, which permits use, distribution and reproduction in any medium, provided the original work is properly cited. |
Keywords: | 19F NMR; DFT calculations; enzyme mechanisms; GTPases; RhoA/RhoGAP |
Dates: |
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Institution: | The University of Sheffield |
Academic Units: | The University of Sheffield > Faculty of Science (Sheffield) > School of Biosciences (Sheffield) > Department of Molecular Biology and Biotechnology (Sheffield) |
Depositing User: | Symplectic Sheffield |
Date Deposited: | 19 Jul 2017 13:45 |
Last Modified: | 03 Aug 2023 11:41 |
Status: | Published |
Publisher: | Wiley |
Refereed: | Yes |
Identification Number: | 10.1002/anie.201703074 |
Open Archives Initiative ID (OAI ID): | oai:eprints.whiterose.ac.uk:119149 |
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