Agonist regulation of D-2 dopamine receptor/G protein interaction - Evidence for agonist selection of G protein subtype

The D-2 dopamine receptor has been expressed in Sf21. insect cells together with the G proteins G(o) and G(i2), using the baculovirus system. Expressi on levels of receptor and G protein (alpha, beta, and gamma subunits) in th e two preparations were similar as shown by binding of [H-3]spiperone and q uantitative Western blot, respectively. For several agonists, binding data were fitted best by a two-binding site model in either preparation, showing interaction of expressed receptor and G protein. For some agonists, bindin g to the higher affinity site was of higher affinity in D-2/G(o) than in th e D-2/G(i2), preparation. Some agonists exhibited binding data that were be st fitted by a two-binding site model in D-2/G(o) and a one-binding site mo del in D-2/G(i2.) Therefore, receptor/G protein interaction seemed to be st ronger in the D-2/G(o) preparation. Agonist stimulation of [S-35]GTP gammaS (guanosine 5'-3-O-(thio)triphosphate) binding in the two preparations also gave evidence for higher affinity D-2/G(o), interaction. In the D-2/G(o) p reparation, agonist stimulation of [S-35]GTP gammaS binding occurred at hig her potency for several agonists, and a higher stimulation (relative to dop amine) was achieved in D-2/G(o) compared with D2/Gi2, Some agonists were ab le to stimulate [S-35]GTP gammaS binding in the D-2/G(o) preparation but no t in D2/Gi2. The extent of D2 receptor selectivity for Go over Gi2 is there fore dependent on the agonist used, and thus agonists may stabilize differe nt conformations of the receptor with different abilities to couple to and activate G proteins.