Tsukamoto T, Sonenberg M
J Clin Invest. 1979 Aug;64(2):534-40. doi: 10.1172/JCI109491.
The effect of catecholamines on membrane-associated protein kinase in the mature human erythrocyte was investigated. Protein kinase activity was assayed after isolation of membranes from intact erythrocytes incubated with and without catecholamines. Activation of the enzyme is expressed as the ratio of the extent of phosphorylation of exogenous protein substrate in the absence to that in the presence of 2.5 microM cyclic AMP (cAMP). The potent beta-adrenergic agonist, (-)isoproterenol (2 microM), (-)epinephrine (10 microM) and (-)norepinephrine (10 microM) stimulated the cAMP-dependent protein kinase in membranes, 38 +/- 7%, 31 +/- 6%, and 30 +/- 6%, respectively. Maximal stimulation of membrane protein kinase by 10 microM (-)epinephrine was obtained approximately equal to 30 min after initiation of the incubation of erythrocytes with the hormone. The concentrations of (-)catecholamines that gave half-maximal stimulation of the membrane protein kinase were 0.17 microM for isoproterenol, 0.35 microM for epinephrine, and 0.63 microM for norepinephrine. The membrane protein kinase response to beta-adrenergic agonists was found to be stereospecific. The stimulation of membrane protein kinase by 10 microM (-)epinephrine was inhibited by the beta-adrenergic antagonist, (-)propranolol with EC50 = 0.60 microM, and the inhibition of agonist stimulation of the cAMP-dependent protein kinase by propranolol was stereospecific. These studies suggest that a functional beta-adrenergic receptor exists in the mature human erythrocyte.
研究了儿茶酚胺对成熟人红细胞中膜相关蛋白激酶的作用。在用和不用儿茶酚胺孵育完整红细胞后分离膜，然后测定蛋白激酶活性。酶的激活表示为在不存在和存在2.5微摩尔环磷酸腺苷（cAMP）时外源蛋白质底物磷酸化程度的比值。强效β-肾上腺素能激动剂（-）异丙肾上腺素（2微摩尔）、（-）肾上腺素（10微摩尔）和（-）去甲肾上腺素（10微摩尔）分别刺激膜中的cAMP依赖性蛋白激酶38±7%、31±6%和30±6%。在用该激素孵育红细胞开始后约30分钟，10微摩尔（-）肾上腺素对膜蛋白激酶的刺激达到最大。使膜蛋白激酶产生半数最大刺激的（-）儿茶酚胺浓度，异丙肾上腺素为0.17微摩尔，肾上腺素为0.35微摩尔，去甲肾上腺素为0.63微摩尔。发现膜蛋白激酶对β-肾上腺素能激动剂的反应具有立体特异性。10微摩尔（-）肾上腺素对膜蛋白激酶的刺激被β-肾上腺素能拮抗剂（-）普萘洛尔抑制，其半数有效浓度（EC50）为0.60微摩尔，并且普萘洛尔对激动剂刺激的cAMP依赖性蛋白激酶的抑制具有立体特异性。这些研究表明成熟人红细胞中存在功能性β-肾上腺素能受体。
