G-protein alpha subunit interaction and guanine nucleotide dissociation inhibitor activity of the dual GoLoco motif protein PCP-2 (Purkinje cell protein-2)

Purkinje cell protein-2 (PCP-2; L7/GPSM4) is a GoLoco motif-containing protein that is specifically expressed in Purkinje and retinal ON bipolar cells. An alternative splice variant of PCP-2 has recently been isolated which contains two GoLoco motifs. Although the second GoLoco motif (GL2) of PCP-2 has been reported to interact with Gα-subunits, a complete biochemical analysis of each individual motif of PCP-2 has not been performed. We demonstrate that the first GoLoco motif (GL1) of PCP-2 is equipotent as a guanine nucleotide dissociation inhibitor (GDI) towards Gα_i1 and Gα_i2, while it has sevenfold lower GDI activity for Gα_i3 and greater than 20-fold lower GDI activity against Gα_o. In contrast we found PCP-2 GL2 to be essentially equipotent as a GDI for all Gα_i subunits, but it had negligible activity toward Gα_o. Using co-immunoprecipitation from COS-7 cells, we found that PCP-2 was only able to interact with Gα_i1 but not Gα_o nor Gα-subunits from other families (Gα_s, Gα_q, or Gα₁₂). Mutational analysis of a non-canonical residue (glycine 24) in human PCP-2 GL1 provided evidence for heterogeneity in mechanisms of Gα_i interactions with GoLoco motifs. Collectively, the data demonstrate that PCP-2 is a comparatively weak GoLoco motif protein that exhibits highest affinity interactions and GDI activity toward Gα_i1, Gα_i2, and Gα_i3 subunits.