Ric-8B is a GTP-dependent G protein alphas guanine nucleotide exchange factor.

ABSTRACT: ric-8 (resistance to inhibitors of cholinesterase 8) genes have positive roles in variegated G protein signaling pathways, including G?(q) and G?(s) regulation of neurotransmission, G?(i)-dependent mitotic spindle positioning during (asymmetric) cell division, and G?(olf)-dependent odorant receptor signaling. Mammalian Ric-8 activities are partitioned between two genes, ric-8A and ric-8B. Ric-8A is a guanine nucleotide exchange factor (GEF) for G?(i)/?(q)/?(12/13) subunits. Ric-8B potentiated G(s) signaling presumably as a G?(s)-class GEF activator, but no demonstration has shown Ric-8B GEF activity. Here, two Ric-8B isoforms were purified and found to be G? subunit GDP release factor/GEFs. In HeLa cells, full-length Ric-8B (Ric-8BFL) bound endogenously expressed G?(s) and lesser amounts of G?(q) and G?(13). Ric-8BFL stimulated guanosine 5'-3-O-(thio)triphosphate (GTP?S) binding to these subunits and G?(olf), whereas the Ric-8B?9 isoform stimulated G?(s short) GTP?S binding only. Michaelis-Menten experiments showed that Ric-8BFL elevated the V(max) of G?(s) steady state GTP hydrolysis and the apparent K(m) values of GTP binding to G?(s) from ?385 nm to an estimated value of ?42 ?M. Directionality of the Ric-8BFL-catalyzed G?(s) exchange reaction was GTP-dependent. At sub-K(m) GTP, Ric-BFL was inhibitory to exchange despite being a rapid GDP release accelerator. Ric-8BFL binds nucleotide-free G?(s) tightly, and near-K(m) GTP levels were required to dissociate the Ric-8B·G? nucleotide-free intermediate to release free Ric-8B and G?-GTP. Ric-8BFL-catalyzed nucleotide exchange probably proceeds in the forward direction to produce G?-GTP in cells.

SUBMITTER: Chan P 

PROVIDER: S-EPMC3103368 | biostudies-literature | 2011 Jun

REPOSITORIES: biostudies-literature