PMID- 28498638
OWN - NLM
STAT- MEDLINE
DCOM- 20190312
LR  - 20220129
IS  - 1521-3773 (Electronic)
IS  - 1433-7851 (Print)
IS  - 1433-7851 (Linking)
VI  - 56
IP  - 33
DP  - 2017 Aug 7
TI  - Assessing the Influence of Mutation on GTPase Transition States by Using X-ray 
      Crystallography, (19) F NMR, and DFT Approaches.
PG  - 9732-9735
LID - 10.1002/anie.201703074 [doi]
AB  - We report X-ray crystallographic and (19) F NMR studies of the G-protein RhoA 
      complexed with MgF(3)(-) , 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 (19) F 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 O(3G) and thence to P(G) . 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 
      RhoGAP(Arg85'Ala) relative to wild-type (WT) RhoGAP. This study demonstrates that 
      (19) F NMR measurements, in combination with X-ray crystallography and DFT 
      calculations, can reliably dissect the response of small GTPases to site-specific 
      modifications.
CI  - (c) 2017 The Authors. Published by Wiley-VCH Verlag GmbH & Co. KGaA.
FAU - Jin, Yi
AU  - Jin Y
AUID- ORCID: 0000-0002-6927-4371
AD  - Department of Molecular Biology and Biotechnology, Krebs Institute, University of 
      Sheffield, Sheffield, S10 2TN, UK.
AD  - School of Chemistry, Cardiff University, Cardiff, CF10 3AT, UK.
FAU - Molt, Robert W Jr
AU  - Molt RW Jr
AD  - School of Chemistry, Cardiff University, Cardiff, CF10 3AT, UK.
AD  - Department of Biochemistry and Molecular Biology, Indiana University School of 
      Medicine, Indianapolis, IN, 46202, USA.
AD  - ENSCO, Inc., Melbourne, FL, 32940, USA.
FAU - Pellegrini, Erika
AU  - Pellegrini E
AD  - Structural Biology Group, ESRF-The European Synchrotron, CS40220, 38043, 
      Grenoble, Cedex 9, France.
FAU - Cliff, Matthew J
AU  - Cliff MJ
AD  - Manchester Institute of Biotechnology, Manchester, M1 7DN, UK.
FAU - Bowler, Matthew W
AU  - Bowler MW
AD  - Structural Biology Group, ESRF-The European Synchrotron, CS40220, 38043, 
      Grenoble, Cedex 9, France.
AD  - European Molecular Biology Laboratory, Grenoble Outstation CS90181, 38042, 
      Grenoble, Cedex 9, France.
FAU - Richards, Nigel G J
AU  - Richards NGJ
AD  - School of Chemistry, Cardiff University, Cardiff, CF10 3AT, UK.
FAU - Blackburn, G Michael
AU  - Blackburn GM
AD  - Department of Molecular Biology and Biotechnology, Krebs Institute, University of 
      Sheffield, Sheffield, S10 2TN, UK.
FAU - Waltho, Jonathan P
AU  - Waltho JP
AD  - Department of Molecular Biology and Biotechnology, Krebs Institute, University of 
      Sheffield, Sheffield, S10 2TN, UK.
AD  - Manchester Institute of Biotechnology, Manchester, M1 7DN, UK.
LA  - eng
GR  - BB/E017541/1/BB_/Biotechnology and Biological Sciences Research Council/United 
      Kingdom
PT  - Journal Article
PT  - Research Support, Non-U.S. Gov't
DEP - 20170524
PL  - Germany
TA  - Angew Chem Int Ed Engl
JT  - Angewandte Chemie (International ed. in English)
JID - 0370543
RN  - 284SYP0193 (Fluorine)
RN  - EC 3.6.1.- (GTP Phosphohydrolases)
SB  - IM
MH  - Crystallography, X-Ray
MH  - *Density Functional Theory
MH  - Fluorine/chemistry
MH  - GTP Phosphohydrolases/chemistry/*genetics/metabolism
MH  - Magnetic Resonance Spectroscopy
MH  - Models, Molecular
MH  - Mutation
PMC - PMC5575484
OTO - NOTNLM
OT  - 19F NMR
OT  - DFT calculations
OT  - GTPases
OT  - RhoA/RhoGAP
OT  - enzyme mechanisms
EDAT- 2017/05/13 06:00
MHDA- 2019/03/13 06:00
PMCR- 2017/08/30
CRDT- 2017/05/13 06:00
PHST- 2017/03/27 00:00 [received]
PHST- 2017/05/13 06:00 [pubmed]
PHST- 2019/03/13 06:00 [medline]
PHST- 2017/05/13 06:00 [entrez]
PHST- 2017/08/30 00:00 [pmc-release]
AID - ANIE201703074 [pii]
AID - 10.1002/anie.201703074 [doi]
PST - ppublish
SO  - Angew Chem Int Ed Engl. 2017 Aug 7;56(33):9732-9735. doi: 10.1002/anie.201703074. 
      Epub 2017 May 24.