Regulation of cholesterol 7 alpha-hydroxylase in the liver. Purification of cholesterol 7 alpha-hydroxylase and the immunochemical evidence for the induction of cholesterol 7 alpha-hydroxylase by cholestyramine and circadian rhythm.
Regulation of cholesterol 7 alpha-hydroxylase in the liver. Purification of cholesterol 7 alpha-hydroxylase and the immunochemical evidence for the induction of cholesterol 7 alpha-hydroxylase by cholestyramine and circadian rhythm.
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DOI：
10.1016/s0021-9258(19)39677-2
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发表时间：
1990-03
期刊：
影响因子：
--
通讯作者：
J. Chiang;W. F. Miller;G. Lin
中科院分区：
文献类型：
--
作者：
J. Chiang;W. F. Miller;G. Lin
使用阴离子交换高效液相色谱从考来烯胺处理的雌性大鼠的肝微粒体中纯化两种胆固醇7α-羟化酶同工酶。这两种细胞色素 P-450 同工酶在电泳迁移率、免疫交叉反应性和 Vmax 方面相似，但在胆固醇 Km、周转数和电荷方面有所不同。在兔体内产生了针对主要同工酶的抗体。该抗体特异性抑制微粒体胆固醇 7 α-羟化酶活性。微粒体多肽的免疫印迹表明，用补充考来烯胺的饮食治疗大鼠后，微粒体胆固醇7α-羟化酶水平与胆固醇7α-羟化酶活性平行增加。从饮食中去除考来烯胺后，胆固醇 7 α-羟化酶活性和酶水平立即急剧降低。在肾脏、心脏和肺的微粒体中也检测到胆固醇 7 α-羟化酶活性，其水平约为肝脏中的 7-27%。 3-甲基胆蒽处理诱导胆固醇 7 α-羟化酶活性和酶水平。相比之下，孕烯醇酮-16α-甲腈或地塞米松治疗极大地抑制了大鼠的酶和活性。维持在反向光周期下的大鼠肝微粒体中胆固醇7α-羟化酶水平比正常光周期下高2-3倍。在遗传性肥胖的 Zucker 大鼠中，胆固醇 7 α-羟化酶活性和酶水平对光周期的变化没有反应，然而，通过考来烯胺治疗将其诱导至与瘦大鼠相同的水平。该研究首次提供直接证据证明微粒体胆固醇7α-羟化酶活性的胆汁酸反馈调节和昼夜节律涉及胆固醇7α-羟化酶水平的诱导。
Two cholesterol 7 alpha-hydroxylase isozymes were purified from liver microsomes of cholestyramine-treated female rats by using anion exchange high performance liquid chromatography. These two cytochrome P-450 isozymes were similar in electrophoretic mobility, immunocross-reactivity, and Vmax but differed in Km for cholesterol, turnover number, and charges. Antibody against the major isozyme was raised in rabbit. This antibody specifically inhibited microsomal cholesterol 7 alpha-hydroxylase activity. Immunoblot of microsomal polypeptides indicated that microsomal cholesterol 7 alpha-hydroxylase enzyme levels were increased in parallel with cholesterol 7 alpha-hydroxylase activity upon the treatment of rats with diet supplemented with cholestyramine. Both cholesterol 7 alpha-hydroxylase activity and enzyme levels were drastically reduced immediately after the removal of cholestyramine from the diet. Cholesterol 7 alpha-hydroxylase activity was also detected in the microsomes of kidney, heart, and lung in about 7-27% of the level found in the liver. 3-Methylcholanthrene treatment induced cholesterol 7 alpha-hydroxylase activity and enzyme level. In contrast, pregnenolone-16 alpha-carbonitrile or dexamethasone treatment greatly depressed enzyme and activity in rats. Cholesterol 7 alpha-hydroxylase enzyme level was 2-3-fold higher in liver microsomes of rats maintained under the reversed light cycle than under the normal light cycle. In genetically obese Zucker rats, cholesterol 7 alpha-hydroxylase activity and enzyme level did not respond to the change in the light cycle, however, were induced to the same levels as in the lean rats by cholestyramine treatment. This study provided the first direct evidence that the bile acid feedback regulation and circadian rhythm of microsomal cholesterol 7 alpha-hydroxylase activity involved the induction of cholesterol 7 alpha-hydroxylase enzyme level.