@article{b7fbcea02b394c6582e6a95bb2f7d47c,
title = "Tyrosine phosphorylation switching of a G protein",
abstract = "Heterotrimeric G protein complexes are molecular switches relaying extracellular signals sensed by G protein-coupled receptors (GPCRs) to downstream targets in the cytoplasm, which effect cellular responses. In the plant heterotrimeric GTPase cycle, GTP hydrolysis, rather than nucleotide exchange, is the rate-limiting reaction and is accelerated by a receptor-like regulator of G signaling (RGS) protein. We hypothesized that post-translational modification of the G subunit in the G protein complex regulates the RGS-dependent GTPase cycle. Our structural analyses identified an invariant phosphorylated tyrosine residue (Tyr166 in the Arabidopsis G subunit AtGPA1) located in the intramolecular domain interface where nucleotide binding and hydrolysis occur. We also identified a receptor-like kinase that phosphorylates AtGPA1 in a Tyr166-dependent manner. Discrete molecular dynamics simulations predicted that phosphorylated Tyr166 forms a salt bridge in this interface and potentially affects the RGS protein-accelerated GTPase cycle. Using a Tyr166 phosphomimetic substitution, we found that the cognate RGS protein binds more tightly to the GDP-bound G substrate, consequently reducing its ability to accelerate GTPase activity. In conclusion, we propose that phosphorylation of Tyr166 in AtGPA1 changes the binding pattern with AtRGS1 and thereby attenuates the steady-state rate of the GTPase cycle. We coin this newly identified mechanism “substrate phosphoswitching.”.",
author = "Bo Li and Meral Tunc-Ozdemir and Daisuke Urano and Haiyan Jia and Werth, {Emily G.} and Mowrey, {David D.} and Hicks, {Leslie M.} and Dokholyan, {Nikolay V.} and Torres, {Matthew P.} and Jones, {Alan M.}",
note = "Funding Information: This work was supported by NIGMS, National Institutes of Health (NIH), Grant R01GM065989 and National Science Foundation (NSF) Grant MCB-1713880 (to A. M. J.); NSF Grant MCB-1552522 (to L. M. H); NIGMS, NIH, Grants R01GM080742, R01GM11401, R01GM064803, and 1R01GM123247 (to N. V. D.); and NIH Grant R01 GM117400 (to M. P. T.). The plant-based experimental part of this project was supported by the Division of Chemi-cal Sciences, Geosciences, and Biosciences, Office of Basic Energy Sciences of the United States Department of Energy through Grant DE-FG02-05er15671 (to A. M. J.). This work was also supported by the NCI, NIH, Grant P30CA016086. The authors declare that they have no conflicts of interest with the contents of this article. The content is solely the responsibility of the authors and does not necessarily represent the official views of the National Institutes of Health. Funding Information: Acknowledgments—We are grateful to Dr. Stephen R. Sprang (University of Montana) for insightful comments; Dr. Henrik Dohlman (University of North Carolina, Chapel Hill, NC) for sharing equipment; and Kevin Knight, Jing Yang, and Catherine Jones for valuable technical assistance. This research is based in part upon work conducted using the UNC Michael Hooker Proteomics Center, which is supported in part by NCI, National Institutes of Health, Grant CA016086 (to the Lineberger Comprehensive Cancer Center). Funding Information: This work was supported by NIGMS, National Institutes of Health (NIH), Grant R01GM065989 and National Science Foundation (NSF) Grant MCB-1713880 (to A. M. J.); NSF Grant MCB-1552522 (to L. M. H); NIGMS, NIH, Grants R01GM080742, R01GM11401, R01GM064803, and 1R01GM123247 (to N. V. D.); and NIH Grant R01 GM117400 (to M. P. T.). The plant-based experimental part of this project was supported by the Division of Chemical Sciences, Geosciences, and Biosciences, Office of Basic Energy Sciences of the United States Department of Energy through Grant DE-FG02-05er15671 (to A. M. J.). This work was also supported by the NCI, NIH, Grant P30CA016086. The authors declare that they have no conflicts of interest with the contents of this article. The content is solely the responsibility of the authors and does not necessarily represent the official views of the National Institutes of Health. Publisher Copyright: {\textcopyright} 2018 by The American Society for Biochemistry and Molecular Biology, Inc. Published in the U.S.A.",
year = "2018",
month = mar,
day = "30",
doi = "10.1074/jbc.RA117.000163",
language = "English (US)",
volume = "293",
pages = "4752--4766",
journal = "Journal of Biological Chemistry",
issn = "0021-9258",
publisher = "American Society for Biochemistry and Molecular Biology Inc.",
number = "13",
}