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. Expression levels of receptor and G protein (alpha, beta, and gamma subunits) in the two preparations were similar as shown by binding of [(3)H]spiperone and quantitative 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, binding to the higher affinity site was of higher affinity in D(2)/G(o) than in the D(2)/G(i2) preparation. Some agonists exhibited binding data that were best fitted by a two-binding site model in D(2)/G(o) and a one-binding site model in D(2)/G(i2). Therefore, receptor/G protein interaction seemed to be stronger in the D(2)/G(o) preparation. Agonist stimulation of [(35)S]GTP gamma S (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) preparation, agonist stimulation of [(35)S]GTP gamma S binding occurred at higher potency for several agonists, and a higher stimulation (relative to dopamine) was achieved in D(2)/G(o) compared with D(2)/G(i2). Some agonists were able to stimulate [(35)S]GTP gamma S binding in the D(2)/G(o) preparation but not in D(2)/G(i2). The extent of D(2) receptor selectivity for G(o) over G(i2) is therefore dependent on the agonist used, and thus agonists may stabilize different conformations of the receptor with different abilities to couple to and activate G proteins.
Cordeaux
Y
Y
School of Animal and Microbial Sciences, University of Reading, Whiteknights, Reading, Berkshire RG6 6AJ, United Kingdom.
Nickolls
S A
SA
Flood
L A
LA
Graber
S G
SG
Strange
P G
PG
D000818:Animals; D016367:Baculoviridae; D001667:Binding, Competitive; D016466:CHO Cells; D002460:Cell Line; D002462:Cell Membrane / Q000378:metabolism; D006224:Cricetinae; D018491:Dopamine Agonists / Q000493:pharmacokinetics / Q000494:pharmacology*; D018492:Dopamine Antagonists / Q000493:pharmacokinetics / Q000494:pharmacology*; D019206:GTP-Binding Protein alpha Subunits, Gi-Go / Q000378:metabolism*; D016244:Guanosine 5'-O-(3-Thiotriphosphate) / Q000378:metabolism*; D020962:Heterotrimeric GTP-Binding Proteins / Q000378:metabolism*; D007700:Kinetics; D011869:Radioligand Assay; D051381:Rats; D017448:Receptors, Dopamine D2 / Q000187:drug effects / Q000235:genetics / Q000502:physiology*; D011994:Recombinant Proteins / Q000187:drug effects / Q000378:metabolism; D013134:Spiperone / Q000493:pharmacokinetics*; D018411:Spodoptera; D013462:Sulfur Radioisotopes; D014162:Transfection; D014316:Tritium
D018491:Dopamine Agonists; D018492:Dopamine Antagonists; D017448:Receptors, Dopamine D2; D011994:Recombinant Proteins; D013462:Sulfur Radioisotopes; D014316:Tritium; D016244:Guanosine 5'-O-(3-Thiotriphosphate); D013134:Spiperone; D019206:GTP-Binding Protein alpha Subunits, Gi-Go; D020962:Heterotrimeric GTP-Binding Proteins