Regulation of the avidity of ternary complexes containing the human 5-HT(1A) receptor by mutation of a receptor contact site on the interacting G protein alpha subunit.
1 Fusion proteins were constructed between the human 5-HT(1A) receptor and pertussis toxin-resistant forms of both G(i1)alpha and G(o1)alpha mutated at residue(351) from cysteine to either glycine or isoleucine. Each of these was expressed stably in HEK293 cells. 2 Increasing concentrations of GDP inhibited binding of the agonist [(3)H]-8-OH-DPAT but not the antagonist [(3)H]-MPPF to each construct. 3 The IC(50) for GDP was greater for constructs containing isoleucine at residue(351) of the G proteins compared to those with glycine at this position. 4 The G protein antagonist suramin had similar effects to GDP on the binding of [(3)H]-8-OH-DPAT. 5 The proportion of 5-HT(1A) receptor binding sites detected by [(3)H]-MPPF that displayed high affinity for 8-OH-DPAT was significantly greater when the interacting G protein contained isoleucine rather than glycine at residue(351). 6 The 5-HT(1A) receptor displayed similar avidity of interaction with G(i1)alpha and G(o1)alpha. 7 These results indicate that a higher avidity ternary complex is formed between 8-OH-DPAT, the 5-HT(1A) receptor and G proteins when isoleucine rather than glycine is located at residue(351) of the interacting G protein.
Welsby
Philip J
PJ
Molecular Pharmacology Group, Division of Biochemistry and Molecular Biology, Institute of Biomedical and Life Sciences, University of Glasgow, Glasgow G12 8QQ.
Carr
I Craig
IC
Wilkinson
Graeme
G
Milligan
Graeme
G
D017371:8-Hydroxy-2-(di-n-propylamino)tetralin / Q000494:pharmacology; D000262:Adenylyl Cyclases / Q000378:metabolism; D000631:Aminopyridines / Q000494:pharmacology; D002460:Cell Line; D005998:Glycine / Q000235:genetics; D006153:Guanosine Diphosphate / Q000494:pharmacology; D020962:Heterotrimeric GTP-Binding Proteins / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism*; D006801:Humans; D007532:Isoleucine / Q000235:genetics; D008024:Ligands; D009154:Mutation; D010879:Piperazines / Q000494:pharmacology; D011869:Radioligand Assay; D011985:Receptors, Serotonin / Q000235:genetics / Q000378:metabolism*; D044263:Receptors, Serotonin, 5-HT1; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism*; D017366:Serotonin Receptor Agonists / Q000494:pharmacology; D013498:Suramin / Q000494:pharmacology
D000631:Aminopyridines; D008024:Ligands; D010879:Piperazines; D011985:Receptors, Serotonin; D044263:Receptors, Serotonin, 5-HT1; D011993:Recombinant Fusion Proteins; D017366:Serotonin Receptor Agonists; D007532:Isoleucine; D006153:Guanosine Diphosphate; D013498:Suramin; D017371:8-Hydroxy-2-(di-n-propylamino)tetralin; C099463:4-(2' methoxyphenyl)-1-(2'-(N-(2''-pyridinyl)-4-fluorobenzamido)ethyl)piperazine; D020962:Heterotrimeric GTP-Binding Proteins; D000262:Adenylyl Cyclases; D005998:Glycine