Inhibition of GDP/GTP exchange on G alpha subunits by proteins containing G-protein regulatory motifs

A novel G alpha binding consensus sequence, termed G-protein regulatory (Ci PR) or GoLoco motif, has been identified in a growing number of proteins, w hich are thought to modulate G-protein signaling. Alternative roles of GPR proteins: as nucleotide exchange factors or as GDP dissociation inhibitors for G alpha have been proposed. We investigated the modulation of the GDP/G TP exchange of Gi alpha (1), Go alpha, and Gs alpha by three proteins conta ining CPR motifs (GPR proteins), LGN-585-642, Pcp2, and RapIGAPII-23-131, t o elucidate the mechanisms of GPR protein function. The GPR proteins displa yed similar patterns of interaction with Gi alpha with the following order of affinities: Gi alpha (1)GDP much greater than Gi alpha (1)GDPAlF(4)(-) g reater than or equal to Gi alpha (1)-GTP gammaS. No detectable binding of t he GPR proteins to Gs alpha was observed, LGN-585-642, Pcp2. and RapIGAPII- 23-131 inhibited the rates of spontaneous GTP gammaS binding and blocked GD P release from Ci alpha (1) and Go alpha. The inhibitory effects of the GPR proteins on Gi alpha (1) were significantly more potent, indicating that G i might be a preferred target for these modulators. Our results suggest tha t GPR proteins are potent GDP dissociation inhibitors for Ci alpha -like G alpha subunits in vitro, and in this capacity they may inhibit GPCR/ Gi pro tein signaling in vivo.