The alpha-helical domain of Galphat determines specific interaction with regulator of G protein signaling 9.

Published

Journal Article

RGS proteins (regulators of G protein signaling) are potent accelerators of the intrinsic GTPase activity of G protein alpha subunits (GAPs), thus controlling the response kinetics of a variety of cell signaling processes. Most RGS domains that have been studied have relatively little GTPase activating specificity especially for G proteins within the Gi subfamily. Retinal RGS9 is unique in its ability to act synergistically with a downstream effector cGMP phosphodiesterase to stimulate the GTPase activity of the alpha subunit of transducin, Galphat. Here we report another unique property of RGS9: high specificity for Galphat. The core (RGS) domain of RGS9 (RGS9) stimulates Galphat GTPase activity by 10-fold and Galphai1 GTPase activity by only 2-fold at a concentration of 10 microM. Using chimeric Galphat/Galphai1 subunits we demonstrated that the alpha-helical domain of Galphat imparts this specificity. The functional effects of RGS9 were well correlated with its affinity for activated Galpha subunits as measured by a change in fluorescence of a mutant Galphat (Chi6b) selectively labeled at Cys-210. Kd values for RGS9 complexes with Galphat and Galphai1 calculated from the direct binding and competition experiments were 185 nM and 2 microM, respectively. The gamma subunit of phosphodiesterase increases the GAP activity of RGS9. We demonstrate that this is because of the ability of Pgamma to increase the affinity of RGS9 for Galphat. A distinct, nonoverlapping pattern of RGS and Pgamma interaction with Galphat suggests a unique mechanism of effector-mediated GAP function of the RGS9.

Full Text

Duke Authors

Cited Authors

  • Skiba, NP; Yang, CS; Huang, T; Bae, H; Hamm, HE

Published Date

  • March 26, 1999

Published In

Volume / Issue

  • 274 / 13

Start / End Page

  • 8770 - 8778

PubMed ID

  • 10085118

Pubmed Central ID

  • 10085118

International Standard Serial Number (ISSN)

  • 0021-9258

Digital Object Identifier (DOI)

  • 10.1074/jbc.274.13.8770

Language

  • eng

Conference Location

  • United States