Department of Pharmacology (D.P.K.), Department of Molecular Physiology and Biological Physics (S.A.L., M.P., H.F., M.C., R.F., M.Y.), Cardiovascular Research Center (M.A.M., R.F., M.Y.), Center for Membrane Biology (M.Y.), and Department of Medicine, Division of Cardiovascular Medicine (M.Y.), University of Virginia School of Medicine, Charlottesville, Virginia; the Skaggs School of Pharmacy and Pharmaceutical Sciences, University of California, San Diego, La Jolla, California (F.M.M., R.A.); and the La Jolla Institute for Allergy and Immunology (J.L.), La Jolla, California.
Mol Pharmacol. 2014 Feb;85(2):301-9. doi: 10.1124/mol.113.088682. Epub 2013 Nov 11.
Allosteric enhancers of the adenosine A1 receptor amplify signaling by orthosteric agonists. Allosteric enhancers are appealing drug candidates because their activity requires that the orthosteric site be occupied by an agonist, thereby conferring specificity to stressed or injured tissues that produce adenosine. To explore the mechanism of allosteric enhancer activity, we examined their action on several A1 receptor constructs, including (1) species variants, (2) species chimeras, (3) alanine scanning mutants, and (4) site-specific mutants. These findings were combined with homology modeling of the A1 receptor and in silico screening of an allosteric enhancer library. The binding modes of known docked allosteric enhancers correlated with the known structure-activity relationship, suggesting that these allosteric enhancers bind to a pocket formed by the second extracellular loop, flanked by residues S150 and M162. We propose a model in which this vestibule controls the entry and efflux of agonists from the orthosteric site and agonist binding elicits a conformational change that enables allosteric enhancer binding. This model provides a mechanism for the observations that allosteric enhancers slow the dissociation of orthosteric agonists but not antagonists.
变构增强剂可放大腺苷 A1 受体的信号转导，其作用机制为变构调节。变构增强剂是一种有吸引力的药物候选物，因为它们的活性需要占据变构结合位点的配体是激动剂，这使得变构增强剂对产生腺苷的应激或受损组织具有特异性。为了探索变构增强剂的作用机制，我们检测了它们对几种 A1 受体构建体的作用，包括（1）物种变体，（2）物种嵌合体，（3）丙氨酸扫描突变体，和（4）定点突变体。将这些发现与 A1 受体的同源建模和变构增强剂文库的计算机筛选相结合。已知结合的变构增强剂的结合模式与已知的构效关系相关，这表明这些变构增强剂结合到由第二个细胞外环形成的口袋中，由 S150 和 M162 残基侧翼。我们提出了一个模型，其中这个前庭控制着变构结合位点的激动剂的进入和流出，并且激动剂结合引发构象变化，从而使变构增强剂结合。该模型为以下观察结果提供了一种机制：变构增强剂可减缓变构结合位点的激动剂的解离，但不减缓拮抗剂的解离。
