title: Regulation of 17,20 lyase activity by cytochrome b5 and by serine phosphorylation of P450c17.
abstract: Cytochrome P450c17 catalyzes the 17alpha-hydroxylase activity required for glucocorticoid synthesis and the 17,20 lyase activity required for sex steroid synthesis. Most P450 enzymes have fixed ratios of their various activities, but the ratio of these two activities of P450c17 is regulated post-translationally. We have shown that serine phosphorylation of P450c17 and the allosteric action of cytochrome b5 increase 17,20 lyase activity, but it has not been apparent whether these two post-translational mechanisms interact. Using purified enzyme systems, we now show that the actions of cytochrome b5 are independent of the state of P450c17 phosphorylation. Suppressing cytochrome b5 expression in human adrenal NCI-H295A cells by >85% with RNA interference had no effect on 17alpha-hydroxylase activity but reduced 17,20 lyase activity by 30%. Increasing P450c17 phosphorylation could compensate for this reduced activity. When expressed in bacteria, human P450c17 required either cytochrome b5 or phosphorylation for 17,20 lyase activity. The combination of cytochrome b5 and phosphorylation was not additive. Cytochrome b5 and phosphorylation enhance 17,20 lyase activity independently of each other, probably by increasing the interaction between P450c17 and NADPH-cytochrome P450 oxidoreductase.
authors.affiliation: Department of Pediatrics and The Metabolic Research Unit, University of California San Francisco, San Francisco, California 94143-0978, USA.
authors.forename: Amit V
authors.initials: AV
authors.lastname: Pandey
authors.forename: Walter L
authors.initials: WL
authors.lastname: Miller
mesh_terms: D006907:17-alpha-Hydroxypregnenolone / Q000378:metabolism; D002460:Cell Line; D015786:Cytochromes b5 / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D006801:Humans; D009251:NADPH-Ferrihemoprotein Reductase / Q000235:genetics / Q000378:metabolism; D010766:Phosphorylation; D020033:Protein Isoforms / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism; D012313:RNA / Q000737:chemistry / Q000378:metabolism; D012694:Serine / Q000378:metabolism*; D013254:Steroid 17-alpha-Hydroxylase / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*
chemical_list: D020033:Protein Isoforms; D006907:17-alpha-Hydroxypregnenolone; D012694:Serine; D012313:RNA; D015786:Cytochromes b5; D013254:Steroid 17-alpha-Hydroxylase; D009251:NADPH-Ferrihemoprotein Reductase