The interaction between D-2 dopamine receptors and GTP-binding proteins.
D-2 dopamine receptors were solubilized from porcine striatal membranes with 0.3% sodium cholate/1 M NaCl and separated from the bulk of the guanine nucleotide-binding regulatory proteins (G-proteins) by Ultrogel AcA34 gel filtration chromatography. The partially purified D-2 receptors were reconstituted in phospholipid vesicles with Gi or Go purified from porcine brain. The dissociation constant (Kd) of the D-2 receptors in the reconstituted vesicles for [3H]spiperone binding was 82-89 pm, which was not affected by the presence or absence of G-proteins. The displacement curve for [3H]spiperone/dopamine was analyzed, assuming that there are two populations of binding sites. The Kd values for the binding sites with high affinity for agonists (HAS) and that for the binding sites with low affinity for agonists (LAS) were approximately 1 microM and 100 microM, respectively. The proportion of HAS was 8% when the receptor preparation was reconstituted into phospholipid vesicles without G-proteins, but it increased to 58-64% with increasing G-protein concentrations. The potency of Go was a little higher than that of Gi. The proportion of HAS in the presence of G-proteins decreased to about 11% on addition of GTP. When G-proteins were treated with islet-activating protein, GTP-sensitive HAS were not observed. These results indicate that at least 50% of the partially purified D-2 receptors interact with both Gi and Go.
D000818:Animals; D001665:Binding Sites; D019204:GTP-Binding Proteins / Q000378:metabolism*; D016244:Guanosine 5'-O-(3-Thiotriphosphate); D006160:Guanosine Triphosphate / Q000031:analogs & derivatives / Q000378:metabolism / Q000494:pharmacology; D066298:In Vitro Techniques; D007473:Ion Channels / Q000502:physiology; D037342:Pertussis Toxin; D011954:Receptors, Dopamine / Q000378:metabolism*; D017448:Receptors, Dopamine D2; D013134:Spiperone / Q000378:metabolism; D013552:Swine; D013873:Thionucleotides / Q000378:metabolism; D010566:Virulence Factors, Bordetella / Q000494:pharmacology
D007473:Ion Channels; D011954:Receptors, Dopamine; D017448:Receptors, Dopamine D2; D013873:Thionucleotides; D010566:Virulence Factors, Bordetella; D016244:Guanosine 5'-O-(3-Thiotriphosphate); D013134:Spiperone; D006160:Guanosine Triphosphate; D037342:Pertussis Toxin; D019204:GTP-Binding Proteins