PHYSICOCHEMICAL CHARACTERIZATION OF PHOTOAFFINITY-LABELED ANGIOTENSIN-II RECEPTORS
PHYSICOCHEMICAL CHARACTERIZATION OF PHOTOAFFINITY-LABELED ANGIOTENSIN-II RECEPTORS
复制标题
DOI：
10.1210/mend-1-2-147
复制
发表时间：
1987-02-01
影响因子：
--
通讯作者：
CATT, KJ
中科院分区：
文献类型：
--
作者：
CARSON, MC;HARPER, CML;CATT, KJ
在用放射性光亲和类似物125 I-[Sar 1，（4-N3）Phe 8]-AII共价标记后，分析肾上腺皮质和其他靶组织（包括肝脏、垂体前叶和平滑肌）中血管紧张素II（AII）的特异性受体位点。光反应性AII衍生物保留了肾上腺受体的高亲和力和肾上腺肾小球细胞中的类固醇生成活性。在牛肾上腺皮质，共价标记的AII受体的光反应性类似物特异性抑制[Sar 1]AII的IC 50约为5 nM。在还原条件下标记蛋白质的聚丙烯酰胺凝胶电泳期间，共价AII-受体复合物的Mr为58，000。在非还原条件下，也观察到一个较小的带与Mr为105，000。两个标记的物种，也发现在凝胶渗透色谱的洗涤剂溶解的复合物，与64,000和86000的夫人。可溶性光标记复合物的pI被吸附到麦胚凝集素Sepharose 6 MB上，并可被N-乙酰氨基葡萄糖洗脱。十二烷基硫酸钠-聚丙烯酰胺凝胶电泳测定的几种靶组织中特定AII结合位点的Mrs显示，十二烷基硫酸钠-聚丙烯酰胺凝胶电泳测定的靶组织在种内和种间表现出显著差异。最显著的差异是大鼠肾上腺皮质（79，000）与大鼠肝脏（60，000）和牛肾上腺皮质（58，000）之间的差异。酶促去糖基化后，牛和大鼠肾上腺皮质中存在的主要成分的Mr分别下降了40%和55%，至35，000和34，000，这表明碳水化合物含量的变化有助于个体靶组织中AII受体的物理异质性。
Specific receptor sites for angiotensin II (AII) were analyzed in the adrenal cortex and other target tissues including liver, anterior pituitary gland, and smooth muscle, after covalent labeling with the radioactive photoaffinity analog 125I-[Sar1,(4-N3)Phe8]-AII. The photoreactive AII derivative retained high affinity for adrenal receptors and full steroidogenic activity in adrenal glomerulosa cells. In bovine adrenal cortex, covalent labeling of AII receptors by the photoreactive analog was specifically inhibited by [Sar1]AII with an IC50 of about 5 nM. The Mr of the covalent AII-receptor complex during polyacrylamide gel electrophoresis of the labeled protein under reducing conditions was 58,000. Under nonreducing conditions, a minor band with Mr of 105,000 was also observed. Two labeled species were also found during gel permeation chromatography of the detergent-solubilized complex, with Mrs of 64,000 and 86,000. The pI of the solubilized photolabeled complex was absorbed to wheat germ lectin Sepharose 6MB and could be eluted by N-acetylglucosamine. The Mrs of specific AII-binding sites in several target tissues, determined by sodium dodecyl sulfate-polyacrylamide gel electrophoresis, showed target tissues, determined by sodium dodecyl sulfate-polyacrylamide gel electrophoresis, showed significant differences within and between species. The most striking differences were between rat adrenal cortex (79,000) and both rat liver (60,000) and bovine adrenal cortex (58,000). After enzymatic deglycosylation, the Mr of the major component present in the bovine and rat adrenal cortex decreased by 40% and 55% to 35,000 and 34,000, respectively, suggesting that variations in carbohydrate content contribute to the physical heterogeneity of AII receptors in individual target tissues.