Assessing the Influence of Mutation on GTPase Transition States by Using X-ray Crystallography, 19 F NMR, and DFT Approaches

Angewandte Chemie
Yi JinJonathan P Waltho

Abstract

We report X-ray crystallographic and 19 F 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 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 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 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.

References

Jan 1, 1995·Nature Structural Biology·R Hilgenfeld
Dec 10, 1998·Nature Structural Biology·N NassarR A Cerione
Apr 3, 2002·Chemistry & Biology·Debbie L GrahamJohn F Eccleston
Sep 23, 2006·Proceedings of the National Academy of Sciences of the United States of America·Nicola J BaxterNicholas H Williams
Nov 21, 2007·FEBS Letters·Henrik te HeesenJürgen Schlitter
Feb 19, 2010·Proceedings of the National Academy of Sciences of the United States of America·Nicola J BaxterJonathan P Waltho
Apr 20, 2010·Journal of the American Chemical Society·Matthew J CliffJonathan P Waltho
Feb 26, 2011·Journal of the American Chemical Society·Liu XiaoxiaJonathan P Waltho
Apr 26, 2011·Annual Review of Biochemistry·Jonathan K LassilaDaniel Herschlag
Apr 17, 2012·Proceedings of the National Academy of Sciences of the United States of America·Joanna L GriffinJonathan P Waltho
Sep 6, 2012·Proceedings of the National Academy of Sciences of the United States of America·Till RudackKlaus Gerwert
Jan 11, 2013·Physiological Reviews·Jacqueline Cherfils, Mahel Zeghouf
Mar 4, 2015·Archives of Biochemistry and Biophysics·Alexandra T P CarvalhoShina C L Kamerlin
Sep 17, 2015·The Journal of Physical Chemistry. B·Maria G KhrenovaAlexander V Nemukhin
Nov 20, 2016·Angewandte Chemie·Yi JinG Michael Blackburn
Dec 3, 2016·Proceedings of the National Academy of Sciences of the United States of America·Daniel MannCarsten Kötting
Sep 25, 2017·Chemistry : a European Journal·Eric A MarroRebekka S Klausen

❮ Previous
Next ❯

Citations

Sep 8, 2021·Nature Communications·Alexander A MalärThomas Wiegand

❮ Previous
Next ❯

Methods Mentioned

BETA
X‐ray
GTPase
NMR
GTPases

Related Concepts

Trending Feeds

COVID-19

Coronaviruses encompass a large family of viruses that cause the common cold as well as more serious diseases, such as the ongoing outbreak of coronavirus disease 2019 (COVID-19; formally known as 2019-nCoV). Coronaviruses can spread from animals to humans; symptoms include fever, cough, shortness of breath, and breathing difficulties; in more severe cases, infection can lead to death. This feed covers recent research on COVID-19.

Blastomycosis

Blastomycosis fungal infections spread through inhaling Blastomyces dermatitidis spores. Discover the latest research on blastomycosis fungal infections here.

Nuclear Pore Complex in ALS/FTD

Alterations in nucleocytoplasmic transport, controlled by the nuclear pore complex, may be involved in the pathomechanism underlying multiple neurodegenerative diseases including Amyotrophic Lateral Sclerosis and Frontotemporal Dementia. Here is the latest research on the nuclear pore complex in ALS and FTD.

Applications of Molecular Barcoding

The concept of molecular barcoding is that each original DNA or RNA molecule is attached to a unique sequence barcode. Sequence reads having different barcodes represent different original molecules, while sequence reads having the same barcode are results of PCR duplication from one original molecule. Discover the latest research on molecular barcoding here.

Chronic Fatigue Syndrome

Chronic fatigue syndrome is a disease characterized by unexplained disabling fatigue; the pathology of which is incompletely understood. Discover the latest research on chronic fatigue syndrome here.

Evolution of Pluripotency

Pluripotency refers to the ability of a cell to develop into three primary germ cell layers of the embryo. This feed focuses on the mechanisms that underlie the evolution of pluripotency. Here is the latest research.

Position Effect Variegation

Position Effect Variagation occurs when a gene is inactivated due to its positioning near heterochromatic regions within a chromosome. Discover the latest research on Position Effect Variagation here.

STING Receptor Agonists

Stimulator of IFN genes (STING) are a group of transmembrane proteins that are involved in the induction of type I interferon that is important in the innate immune response. The stimulation of STING has been an active area of research in the treatment of cancer and infectious diseases. Here is the latest research on STING receptor agonists.

Microbicide

Microbicides are products that can be applied to vaginal or rectal mucosal surfaces with the goal of preventing, or at least significantly reducing, the transmission of sexually transmitted infections. Here is the latest research on microbicides.