Yeung S M, Perez-Reyes E, Cooper D M
Department of Pharmacology, University of Colorado Health Sciences Center, Denver 80262.
Biochem J. 1987 Dec 15;248(3):635-42. doi: 10.1042/bj2480635.
Adenosine Ri receptors and inhibitory guanine-nucleotide-regulatory components were solubilized from rat cerebral-cortical membranes with sodium cholate. (-)-N6-Phenylisopropyl[2,8-3H]adenosine [( 3H]PIA) binds with high affinity to the soluble receptors, which retain the pharmacological specificity of adenosine Ri receptors observed in membranes. The binding is regulated by bivalent cations and guanine nucleotides. Bivalent cations increase [3H]PIA binding by increasing both the affinity and the apparent number of receptors. Guanine nucleotides decrease agonist binding by increasing the dissociation of the ligand-receptor complex. Adenosine agonists stabilize the high-affinity form of the soluble receptor. The hydrodynamic properties of the adenosine receptor were determined with cholate extracts of membranes that were treated with [3H]PIA. Sucrose-gradient-centrifugation analysis indicates that the receptor has a sedimentation coefficient of 7.7 S. The receptor is eluted from Sepharose 6B columns with an apparent Stokes radius of 7.2 nm. Labelling of either sucrose-gradient or gel-filtration-column fractions with pertussis toxin and [32P]-NAD+ reveals that both the 41,000- and 39,000-Mr substrates overlap with the receptor activity. These studies suggest that the high-affinity adenosine-receptor-binding activity in the cholate extract represents a stable R1-N complex.
用胆酸钠从大鼠大脑皮层膜中溶解出腺苷R1受体和抑制性鸟嘌呤核苷酸调节成分。（-）-N6-苯异丙基[2,8-3H]腺苷[（3H]PIA）与可溶性受体高亲和力结合，该受体保留了在膜中观察到的腺苷R1受体的药理学特异性。这种结合受二价阳离子和鸟嘌呤核苷酸的调节。二价阳离子通过增加受体的亲和力和表观数量来增加[3H]PIA的结合。鸟嘌呤核苷酸通过增加配体-受体复合物的解离来降低激动剂结合。腺苷激动剂稳定可溶性受体的高亲和力形式。用[3H]PIA处理的膜的胆酸盐提取物测定腺苷受体的流体动力学性质。蔗糖梯度离心分析表明该受体的沉降系数为7.7 S。该受体从琼脂糖6B柱上洗脱，表观斯托克斯半径为7.2 nm。用百日咳毒素和[32P]-NAD+对蔗糖梯度或凝胶过滤柱级分进行标记，结果显示41000和39000 Mr的底物都与受体活性重叠。这些研究表明胆酸盐提取物中的高亲和力腺苷受体结合活性代表一种稳定的R1-N复合物。
