26-hydroxylation Of C27-steroids Research Articles

25-Hydroxylation of vitamin D3 by a cytochrome P-450 from rabbit liver mitochondria.

A cytochrome P-450 catalysing 25-hydroxylation of vitamin D3 was purified from liver mitochondria of untreated rabbits. The enzyme fraction contained 9 nmol of cytochrome P-450/mg of protein and showed only one protein band with an apparent Mr of 52,000 upon SDS/polyacrylamide-gel electrophoresis. The preparation showed a single protein spot with an apparent isoelectric point of 7.8 and an Mr of approx. 52,000 upon two-dimensional isoelectric-focusing-polyacrylamide-gel electrophoresis. The purified cytochrome P-450 catalysed 25-hydroxylation of vitamin D3 up to 5000 times more efficiently than did the mitochondria. The cytochrome P-450 required both ferredoxin and ferredoxin reductase for catalytic activity. Microsomal NADPH-cytochrome P-450 reductase could not replace ferredoxin and ferredoxin reductase. The cytochrome P-450 catalysed, in addition to 25-hydroxylation of vitamin D3, the 25-hydroxylation of 1 alpha-hydroxyvitamin D3 and the 26-hydroxylation of 5 beta-cholestane-3 alpha, 7 alpha, 12 alpha-triol. The enzyme did not catalyse side-chain cleavage of cholesterol, 11 beta-hydroxylation of deoxycorticosterone, 1 alpha-hydroxylation of 25-hydroxyvitamin D3, hydroxylations of lauric acid and testosterone or demethylation of benzphetamine. The results raise the possibility that the 25-hydroxylation of vitamin D3 and the 26-hydroxylation of C27 steroids are catalysed by the same species of cytochrome P-450 in liver mitochondria. The possible role of the liver mitochondrial cytochrome P-450 in the metabolism of vitamin D3 is discussed.

26-Hydroxylation of C27-steroids by soluble liver mitochondrial cytochrome P-450.

26-Hydroxylation of C27-steroids by soluble liver mitochondrial cytochrome P-450.

Effect of biliary obstruction on 26-hydroxylation of C27-steroids in bile acid synthesis

The effect of biliary obstruction on side chain hydroxylations in the biosynthesis and metabolism of bile acids was studied in the rat. For comparison, several other hydroxylation reactions in bile acid biosynthesis and metabolism were assayed. Biliary obstruction inhibited microsomal 26-hydroxylation of 5beta-cholestane-3alpha,7alpha-diol and microsomal 25- and 26-hydroxylation of 5beta-cholestane-3alpha,7alpha-12alpha-triol. Microsomal 7alpha-hydroxylation of cholesterol and 6beta-hydroxylation of lithocholic acid acid increased significantly, whereas the increase in microsomal 12alpha-hydroxylation of 5beta-cholestane-3alpha,7alpha-diol was less. Mitochondrial 26-hydroxylation of cholesterol, 5-cholestene-3beta,7alpha-diol, and 7alpha-hydroxy-4-cholesten-3-one was stimulated, whereas 26-hydroxylation of 5beta-cholestane-3alpha,7alpha-diol was not affected and that of 5beta-cholestane-3alpha,7alpha,12alpha-triol was markedly inhibited. The results indicate that mitochondrial 26-hydroxylation, particularly of substrates that primarily are precursors of chenodeoxycholic acid, plays a more important role in bile acid biosynthesis under conditions of biliary obstruction than under normal conditions.