Gorbachevskaia L V
Vopr Med Khim. 1978 May-Jun;24(3):423-7.
A method for evaluation of binding constants and of amount of cAMP binding sites in crude tissue extracts was developed. The method is based on equilibrium binding of 3H-cAMP by proteins with subsequent ultrafiltration. Hydrolysis of cAMP and its unspecific sorption by proteins were eliminated under the conditions selected. Rat spleen cytosole contained 3.57 +/- 0.34 pmol of cAMP binding sites per mg of protein with dissociation constant of protein-cAMP complex (1.68 +/- 0.28).10(-8) M. As shown by studies on kinetics, binding constants and specificity of binding, the method permitted to evaluate quantitatively cAMP-dependent protein kinases in crude tissue extracts and to estimate their affinity to cAMP.
开发了一种评估粗组织提取物中结合常数和环磷酸腺苷（cAMP）结合位点数量的方法。该方法基于蛋白质对³H-cAMP的平衡结合以及随后的超滤。在所选择的条件下，消除了cAMP的水解及其被蛋白质的非特异性吸附。大鼠脾脏胞质溶胶每毫克蛋白质含有3.57±0.34皮摩尔的cAMP结合位点，蛋白质-cAMP复合物的解离常数为（1.68±0.28）×10⁻⁸M。动力学研究、结合常数和结合特异性研究表明，该方法能够定量评估粗组织提取物中环磷酸腺苷依赖性蛋白激酶，并估计它们对cAMP的亲和力。
