Biochemical purification and functional analysis of complexes between the G-protein subunit Gβ5 and RGS proteins

D. Scott Witherow, Vladlen Z Slepak

Research output: Contribution to journalArticle

5 Scopus citations

Abstract

Regulator of G-protein signaling (RGS) proteins of the R7 subfamily (RGS6, 7, 9, and 11) contain a unique Gγ-like (GGL) domain that enables their association with the G-protein β subunit Gβ5. The existence of these complexes was demonstrated by their purification from native tissues as well as by reconstitution in vitro. According to pulse-chase analysis, Gβ5 and RGS7 monomers undergo rapid proteolytic degradation in cells, whereas the dimer is stable. Studies of the functional role of Gβ5-RGS dimers using GTPase activity, ion channel, and calcium mobilization assays showed that, similarly to other RGS proteins, they can negatively regulate G-protein-mediated signal transduction. Protein-protein interactions involving the Gβ5-RGS7 complex can be studied in cells using fluorescence resonance energy transfer utilizing Gβ5, RGS, and Gα subunits fused to the cyan and yellow versions of green fluorescent protein.

Original languageEnglish
Pages (from-to)149-162
Number of pages14
JournalMethods in Enzymology
Volume390
DOIs
StatePublished - Oct 27 2004
Externally publishedYes

ASJC Scopus subject areas

  • Biochemistry

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