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Formation of 7α‐ and 7β‐hydroxylated bile acid precursors from 27‐hydroxycholesterol in human liver microsomes and mitochondria
Author(s) -
Shoda Junichi,
Toll Anders,
Axelson Magnus,
Pieper Fritz,
Wikvall Kjell,
Sjövall Jan
Publication year - 1993
Publication title -
hepatology
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 5.488
H-Index - 361
eISSN - 1527-3350
pISSN - 0270-9139
DOI - 10.1002/hep.1840170308
Subject(s) - hydroxylation , microsome , biochemistry , enzyme , chemistry , reductase , cytochrome , mitochondrion , cytochrome p450 , bile acid , metabolism
In a search for enzymes involved in the formation of bile acids from 27‐hydroxycholesterol in humans, the metabolism of this and other side‐chain oxygenated steroids was studied in human liver microsomes and mitochondria. The microsomal fraction contained enzyme(s) catalyzing 7α‐hydroxylation of 27‐hydroxycholesterol and 3β‐hydroxy‐5‐cholestenoic acid, whereas the 7α‐hydroxylation of cholesterol and 3β‐hydroxy‐5‐cholenoic acid was low. Only small amounts of 7β‐hydroxylated products were formed. Purification and subfractionation of microsomal protein yielded a fraction of cytochrome P‐450, which required NADPH and NADPH‐cytochrome P‐450 reductase and catalyzed 7α‐hydroxylation of the side‐chain oxygenated 3β‐hydroxy‐δ 5 ‐C 27 ‐steroids but was inactive toward cholesterol. Added cholesterol did not inhibit the observed enzymatic activity. The results provide evidence that this enzyme is different from cholesterol 7α‐hydroxylase. The mitochondrial fraction contained enzyme(s) that catalyzed an isocitrate‐dependent 7α‐hydroxylation of 3β‐hydroxy‐5‐cholestenoic acid. The activity was much lower with 27‐hydroxycholesterol. The mitochondrial fraction also catalyzed the oxidation of the 27‐hydroxy group and contained a 3β‐hydroxy‐δ 5 ‐steroid dehydrogenase active on 7α‐hydroxylated C 27 ‐steroids. The metabolic end product of the reactions catalyzed by these enzymes was 7α‐hydroxy‐3‐oxo‐4‐cholestenoic acid. A considerable fraction of the 7α‐hydroxy‐δ 5 intermediates was also converted to the corresponding 7β‐hydroxysteroids, probably by way of the 7‐oxosteroids, suggesting the presence of an epimerizing enzyme in the mitochondrial fraction. This study shows that 7α‐hydroxy‐3‐oxo‐4‐cholestenoic acid, which is believed to be an important precursor of chenodeoxycholic acid in human beings, can be formed from 27‐hydroxycholesterol or 3β‐hydroxy‐5‐cholestenoic acid by the action of 7α‐hydroxylases present in the microsomal and mitochondrial fractions. These enzymes, found to be more active in humans and pigs than in rats and rabbits, can catalyze the formation of bile acids by way of pathways that bypass the microsomal cholesterol 7α‐hydroxylase as the rate‐limiting enzyme. (H EPATOLOGY 1993;17:395–403.)