The role of Mg2+ on the formation of the ternary complex between agonist, beta-adrenoceptor, and GS-protein and an interpretation of high and low affinity binding of beta-adrenoceptor agonists.
In the absence of GTP or its analogues, the beta-adrenoceptor agonist (H), the receptor (R) and the stimulatory guanine nucleotide-binding regulatory protein (GS-protein) form a slowly dissociable complex (HRGSGDP). It is presumed that the agonist in this ternary complex is bound to the receptor with high affinity while its binding to the receptor in the HR-complex is of low affinity. We have studied the effect of Mg2+ on the formation of these two complexes by the displacement of (-)-[3H]CGP-12177 with (-)-isoprenaline in cell membranes prepared from guinea-pig lung parenchyma. In the absence of Mg2+, only monophasic displacement curves were obtained with a dissociation constant not differing from that obtained in the presence of Gpp(NH)p and Mg2+. In the presence of Mg2+ and absence of Gpp(NH)p, a shallow binding curve was obtained that fitted a two site model best. One of the dissociation constants agreed well with that obtained in the presence of Gpp(NH)p and Mg2+ but the other one was 27 times smaller. It was concluded that Mg2+ is necessary for the coupling between the HR-complex and the GS-protein. The two dissociation constants obtained in the absence of Gpp(NH)p represent the dissociation constants of the HR- and HRGSGDP-complexes and may correspond to those named KL and KH in the literature. According to our study, there may be no difference in affinity between the agonist and receptor in the HR- and HRGSGDP-complexes. The two dissociation constants obtained in the absence of GTP or its analogues refer to two consecutive reactions.
Preclinical Research and Development Department, Astra Draco AB, Lund, Sweden.
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Johansson
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Persson
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Rosengren
D000318:Adrenergic beta-Agonists / Q000378:metabolism*; D000319:Adrenergic beta-Antagonists / Q000378:metabolism; D000818:Animals; D002462:Cell Membrane / Q000187:drug effects / Q000378:metabolism; D019204:GTP-Binding Proteins / Q000378:metabolism*; D006168:Guinea Pigs; D007545:Isoproterenol / Q000494:pharmacology; D008168:Lung / Q000187:drug effects / Q000378:metabolism; D008274:Magnesium / Q000494:pharmacology*; D008297:Male; D011412:Propanolamines / Q000378:metabolism; D011943:Receptors, Adrenergic, beta / Q000187:drug effects* / Q000378:metabolism
D000318:Adrenergic beta-Agonists; D000319:Adrenergic beta-Antagonists; D011412:Propanolamines; D011943:Receptors, Adrenergic, beta; D019204:GTP-Binding Proteins; D008274:Magnesium; D007545:Isoproterenol; C033394:CGP 12177