Cornett L E, Norris J S
Mol Cell Biochem. 1985 May;67(1):47-53. doi: 10.1007/BF00220985.
In this study, we have used an alpha 1-adrenergic receptor photoaffinity ligand, 2-[4-(4-azido-3-iodo-benzoyl)-piperazin-1-yl]-4-amino-6, 7-dimethoxyquinazoline (125I-APD), to label covalently the alpha 1-adrenergic receptor in a smooth muscle cell line. Our results indicate that in the absence of light, (125I)APD binds reversibly to a site in the DDT1 MF-2 cell membranes having pharmacological characteristics of an alpha 1-adrenergic receptor. Following incorporation of (125I)APD into partially purified membranes a single labeled band of protein with a Mr of 81 000 was visualized by autoradiography following sodium dodecyl sulfate-polyacrylamide gel electrophoresis. Incorporation of (125I)-APD into this band was affected by adrenergic agonists and antagonists in a manner consistent with an alpha 1-adrenergic interaction. Prazosin (alpha 1-selective) blocked incorporation of the label into the Mr = 81 000 protein while yohimbine (alpha 2-selective) did not. Of the adrenergic agonists, (-)-epinephrine and (-)-norepinephrine but not (-)-isoproterenol blocked labeling of the Mr = 81 000 protein. We conclude that the ligand binding site of the DDT1 MF-2 cell alpha 1-adrenergic receptor resides in a Mr = 81 000 protein.
在本研究中，我们使用了一种α1 - 肾上腺素能受体光亲和配体，即2 - [4 - (4 - 叠氮基 - 3 - 碘 - 苯甲酰基) - 哌嗪 - 1 - 基] - 4 - 氨基 - 6,7 - 二甲氧基喹唑啉(125I - APD)，对平滑肌细胞系中的α1 - 肾上腺素能受体进行共价标记。我们的结果表明，在无光条件下，(125I)APD可逆地结合到DDT1 MF - 2细胞膜上具有α1 - 肾上腺素能受体药理学特性的位点。将(125I)APD掺入部分纯化的膜中后，在十二烷基硫酸钠 - 聚丙烯酰胺凝胶电泳后通过放射自显影观察到一条单一的Mr为81 000的标记蛋白带。(125I) - APD掺入该条带受到肾上腺素能激动剂和拮抗剂的影响，其方式与α1 - 肾上腺素能相互作用一致。哌唑嗪(α1选择性)可阻断标记物掺入Mr = 81 000的蛋白中，而育亨宾(α2选择性)则不能。在肾上腺素能激动剂中，(-) - 肾上腺素和(-) - 去甲肾上腺素可阻断Mr = 81 000蛋白的标记，而(-) - 异丙肾上腺素则不能。我们得出结论，DDT1 MF - 2细胞α1 - 肾上腺素能受体的配体结合位点存在于一个Mr = 81 000的蛋白中。
