(19)F NMR and DFT Analysis Reveal Structural and Electronic Transition State Features for RhoA-Catalyzed GTP Hydrolysis

Angewandte Chemie
Yi JinG Michael Blackburn

Abstract

Molecular details for RhoA/GAP catalysis of the hydrolysis of GTP to GDP are poorly understood. We use (19)F NMR chemical shifts in the MgF3(-) transition state analogue (TSA) complex as a spectroscopic reporter to indicate electron distribution for the γ-PO3(-) oxygens in the corresponding TS, implying that oxygen coordinated to Mg has the greatest electron density. This was validated by QM calculations giving a picture of the electronic properties of the transition state (TS) for nucleophilic attack of water on the γ-PO3(-) group based on the structure of a RhoA/GAP-GDP-MgF3(-) TSA complex. The TS model displays a network of 20 hydrogen bonds, including the GAP Arg85' side chain, but neither phosphate torsional strain nor general base catalysis is evident. The nucleophilic water occupies a reactive location different from that in multiple ground state complexes, arising from reorientation of the Gln-63 carboxamide by Arg85' to preclude direct hydrogen bonding from water to the target γ-PO3(-) group.

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Citations

Dec 3, 2016·Proceedings of the National Academy of Sciences of the United States of America·Daniel MannCarsten Kötting
Mar 17, 2017·Topics in Current Chemistry·Yi JinG Michael Blackburn
Nov 20, 2016·Angewandte Chemie·Yi JinG Michael Blackburn
Mar 16, 2017·Acta Crystallographica. Section D, Structural Biology·Heping ZhengWladek Minor
Apr 30, 2017·Chemical Science·Ravi TripathiDominik Marx
Nov 4, 2020·Nature Communications·Henry P WoodJonathan P Waltho
Mar 17, 2020·ACS Omega·Ana Rita CalixtoShina Caroline Lynn Kamerlin
Nov 10, 2018·ACS Central Science·Marten PrießLars V Schäfer
Jun 15, 2019·Journal of the American Chemical Society·Ana R CalixtoShina C L Kamerlin

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Methods Mentioned

BETA
GTPases
NMR

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