Purification of heterotrimeric G protein alpha subunits by GST-Ric-8 association: primary characterization of purified G alpha(olf).

ABSTRACT: Ric-8A and Ric-8B are nonreceptor G protein guanine nucleotide exchange factors that collectively bind the four subfamilies of G protein α subunits. Co-expression of Gα subunits with Ric-8A or Ric-8B in HEK293 cells or insect cells greatly promoted Gα protein expression. We exploited these characteristics of Ric-8 proteins to develop a simplified method for recombinant G protein α subunit purification that was applicable to all Gα subunit classes. The method allowed production of the olfactory adenylyl cyclase stimulatory protein Gα(olf) for the first time and unprecedented yield of Gα(q) and Gα(13). Gα subunits were co-expressed with GST-tagged Ric-8A or Ric-8B in insect cells. GST-Ric-8·Gα complexes were isolated from whole cell detergent lysates with glutathione-Sepharose. Gα subunits w

SUBMITTER: Chan P 

PROVIDER: S-EPMC3024758 | biostudies-literature | 2011 Jan

REPOSITORIES: biostudies-literature