Mahle C D, Wiener H L, Yocca F D, Maayani S
Department of Pharmacology, Mount Sinai School of Medicine, City University of New York, New York.
J Pharmacol Exp Ther. 1992 Dec;263(3):1275-84.
The ternary complex formed between agonist, receptor and guanine nucleotide binding protein and its destabilization by guanine nucleotides (GN) was utilized to study early events in signal transduction, by characterizing the allosteric interactions between agonist and GN binding to the receptor/guanine nucleotide binding protein, G complex for adenosine A1 and 5-hydroxytryptamine1A receptors. The functional interaction between the ternary complex and GTP was examined by assaying adenylyl cyclase activity. Binding of a full adenosine A1 agonist ([3H]-R-(-)-N6-(2-phenylisopropyl)adenosine), and a full [(+-)-[3H]-8-hydroxydipropylaminotetralin] and partial ([3H]-8-[2-[4-(2-methoxyphenyl)-1-piperazinyl]ethyl]-8- azaspirol[4.5]-decane-7,9-dione) 5-hydroxytryptamine1A agonist was examined in relation to the binding of GN. The amount of ternary complex formed depended upon receptor type and drug relative efficacy. The ratio between the drug's EC50 value (adenylyl cyclase) and dissociation constant (binding) was also receptor and drug relative efficacy dependent. 5'-Guanylylimidodiphosphate (100 microM) caused an approximately 50% decrease in the Bmax for all drugs without affecting Kd values. 5'-Guanylylimidodiphosphate and guanosine 5'-O-(3-thiotriphosphate) attenuated [3H]-agonist binding in a concentration-dependent and saturable manner, with IC50 values increased 2- to 6-fold with increasing receptor occupancy. IC50 values were approximately one-tenth lower at the 5-hydroxytryptamine1A receptor than adenosine A1 receptor; similar values were obtained for inhibition of (+-)-[3H]-8-hydroxydipropylaminotetralin and [3H]-8-[2-[4-(2-methoxyphenyl)-1-piperazinyl]ethyl]-8- azaspirol[4.5]-decane-7,9-dione binding, suggesting an independence of agonist efficacy. We propose that the stabilization of the ternary complex by hormone binding, measured by Bmax values, is related to drug-relative efficacy, thus the amount of ternary complex available for destabilization by GN is greater for the more efficacious agonist. This is translated into greater relative efficacy observed in the maximal inhibition of adenylyl cyclase.
激动剂、受体和鸟嘌呤核苷酸结合蛋白之间形成的三元复合物及其被鸟嘌呤核苷酸（GN）破坏稳定性的过程，被用于研究信号转导的早期事件，通过表征激动剂与结合到受体/鸟嘌呤核苷酸结合蛋白（腺苷A1和5-羟色胺1A受体的G复合物）上的GN之间的变构相互作用。通过测定腺苷酸环化酶活性来检测三元复合物与GTP之间的功能相互作用。研究了完全腺苷A1激动剂（[3H]-R-(-)-N6-(2-苯异丙基)腺苷）、完全激动剂[(+-)-[3H]-8-羟基二丙基氨基四氢萘]和部分激动剂（[3H]-8-[2-[4-(2-甲氧基苯基)-1-哌嗪基]乙基]-8-氮杂螺[4.5]-癸烷-7,9-二酮）5-羟色胺1A激动剂的结合与GN结合的关系。形成的三元复合物的量取决于受体类型和药物相对效能。药物的EC50值（腺苷酸环化酶）与解离常数（结合）之间的比值也取决于受体和药物相对效能。5'-鸟苷酰亚胺二磷酸（100 microM）使所有药物的Bmax值降低约50%，而不影响Kd值。5'-鸟苷酰亚胺二磷酸和鸟苷5'-O-(3-硫代三磷酸)以浓度依赖性和饱和性方式减弱[3H]-激动剂结合，随着受体占有率增加，IC50值增加2至6倍。5-羟色胺1A受体的IC50值比腺苷A1受体低约十分之一；对于抑制(+-)-[3H]-8-羟基二丙基氨基四氢萘和[3H]-8-[2-[4-(2-甲氧基苯基)-1-哌嗪基]乙基]-8-氮杂螺[4.5]-癸烷-7,9-二酮结合获得了相似的值，表明激动剂效能具有独立性。我们提出，通过Bmax值测量的激素结合对三元复合物的稳定作用与药物相对效能有关，因此对于更有效的激动剂，可被GN破坏稳定性的三元复合物的量更大。这转化为在腺苷酸环化酶最大抑制中观察到的更大相对效能。
