Reconstitution of a functional beta-adrenergic receptor using cholate and a novel method for its functional assay.
Reconstitution of a functional beta-adrenergic receptor using cholate and a novel method for its functional assay.
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使用胆酸盐重建功能性β-肾上腺素受体及其功能测定的新方法。
DOI：
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发表时间：
1983
期刊：
影响因子：
--
通讯作者：
A. Levitzki
中科院分区：
文献类型：
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作者：
A. Gal;S. Braun;D. Feder;A. Levitzki
纯化的火鸡红细胞膜在增加胆酸盐与蛋白质的比例和盐的存在下溶解，提取高达20%的β-肾上腺素能受体以及腺苷酸环化酶的GTP刺激蛋白（Ns）。在去除胆酸盐后，通过在生物珠上的主动吸收，β-受体和GTP调节蛋白Ns之间的功能性相互作用定量恢复。在l-异丙肾上腺素和p[NH]ppG存在下，受体（R）能够催化Ns活化至其永久活性状态，N的p[NH]ppG，速率常数（kon）与天然膜的速率常数相同。使用聚（乙二醇）-6000重构R/Ns混合物可与SM-2生物珠一样有效地恢复受体结合特性。然而，与SM-2生物珠不同，聚（乙二醇）在恢复R至Ns功能偶联方面不那么有效。在这篇文章中，我们还报告了定量监测N的能力。p[NH]ppG，在Lubrol-PX作为腺苷酸环化酶的催化单元（c）的来源的存在下使用天然火鸡红细胞膜。后一种方法与使用S49 AC-淋巴瘤细胞膜一样有效，但便宜得多。使用这种技术，我们还表明，当NS到C的相互作用是无效的，采用治疗与N-乙基马来酰亚胺，表征R到NS耦合的参数保持不变。
Solubilization of purified turkey erythrocyte membranes at increasing cholate to protein ratios and in the presence of salt, extracts up to 20% of the beta-adrenergic receptor together with the GTP stimulatory protein (Ns) of adenylate cyclase. Upon removal of the cholate, by active absorption on Bio-beads, the functional interaction between the beta-receptor and the GTP regulatory protein Ns is quantitatively restored. The receptor (R) in the presence of l-isoproterenol and p[NH]ppG is able to catalyze the activation of Ns to its permanently active state, N's p[NH]ppG, with a rate constant (kon) identical to that of the native membrane. Reconstitution of the R/Ns mixture using poly(ethyleneglycol)-6000 restores the receptor binding properties as effectively as SM-2 Bio-beads. Unlike SM-2 Bio-beads, however, poly(ethyleneglycol) is not as efficient in restoring the R to Ns functional coupling. In this communication we also report on the ability to monitor quantitatively N's . p[NH]ppG, using native turkey erythrocyte membranes in the presence of Lubrol-PX as the source of the catalytic unit (c) of adenylate cyclase. The latter method is as efficient as using S49 AC- lymphoma cell membranes but much less expensive. Using this technique, we also demonstrate that when the Ns to C interaction is nullified, employing treatment with N-ethylmaleimide, the parameters which characterize R to Ns coupling remain unchanged.
腺苷酸环化酶的调节成分。
DOI：
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发表时间：
1981
期刊：
The Journal of biological chemistry
影响因子：
--
作者：
Hanski,E;Sternweis,PC;Northup,JK;Dromerick,AW;Gilman,AG
通讯作者：
Gilman,AG
β-肾上腺素能受体和腺苷酸环化酶刺激性 GTP 结合蛋白的功能重建。
DOI：
10.1073/pnas.79.23.7228
发表时间：
1982
影响因子：
11.1
作者：
Pedersen,SE;Ross,EM
通讯作者：
Ross,EM
DOI：
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发表时间：
1981
期刊：
The Journal of biological chemistry
影响因子：
--
作者：
Sternweis,PC;Northup,JK;Smigel,MD;Gilman,AG
通讯作者：
Gilman,AG