Journal of Clinical Endocrinology & Metabolism, Vol 77, 98-102, Copyright © 1993 by Endocrine Society

ARTICLES

Progesterone 16 alpha-hydroxylase activity is catalyzed by human cytochrome P450 17 alpha-hydroxylase

P Swart, AC Swart, MR Waterman, RW Estabrook and JI Mason
Cecil H. and Ida Green Center for Reproductive Biology Sciences, Departments of Biochemistry and Obstetrics and Gynecology, University of Texas Southwestern Medical Center, Dallas 75235.

Progesterone and pregnenolone are metabolized to 17 alpha- hydroxysteroids by a cytochrome P450-dependent 17 alpha-hydroxylase (P450c17). The same enzyme can also catalyze the removal of the side- chain of these 17 alpha-hydroxylated steroids to yield androstenedione and dehydroepiandrosterone, respectively. We investigated the metabolism of progesterone by monkey kidney tumor (COS 1) cells transfected with a plasmid vector containing the cDNA encoding the complete amino acid sequence for human cytochrome P450c17. Transfected COS 1 cells converted progesterone to 17 alpha-hydroxyprogesterone as well as 16 alpha-hydroxyprogesterone, but no detectable androstenedione was produced. However, pregnenolone was converted to 17 alpha- hydroxypregnenolone and, ultimately, dehydroepiandrosterone. No 16 alpha-hydroxypregnenolone was produced. The kinetics of progesterone metabolism by the enzyme expressed in COS 1 cells indicated that both 17 alpha- and 16 alpha-hydroxylated products were products were produced from a common active site. Microsomes prepared from fetal adrenal and adult testis converted progesterone to 17 alpha- hydroxyprogesterone as well as 16 alpha-hydroxyprogesterone. No detectable androstenedione was produced by these preparations. Antibodies raised against porcine cytochrome P450c17 inhibited the 17 alpha- and 16 alpha-hydroxylation of progesterone to the same extent when using fetal adrenal microsomes, whereas no inhibition of 21- hydroxylation of progesterone was observed. Similar results were obtained with the imidazole antimycotic agent ketoconazole, which is a preferential cytochrome P450c17 inhibitor. From these results we conclude that human cytochrome P450c17 exhibits marked progesterone 16 alpha-hydroxylase activity in addition to its 17 alpha-hydroxylase function when expressed not only in a heterologous cell expression system but also, importantly, in human steroidogenic cells. Furthermore, the human enzyme has extremely low C-17,20-lyase activity toward progesterone, 17 alpha-hydroxyprogesterone, and 16 alpha- hydroxyprogesterone and fails to convert these to corresponding C19 steroids.

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