MOLECULAR MODELING OF HUMAN P450C17 & P450C21: ADRENARCHE, ANDROGEN

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Cytochrome P450c17 (17-hydroxylase, 17,20-lyase) catalyzes both the 17-hydroxylation and oxidative cleavage of C21 steroids in both human adrenal glands and gonads. These two activities, however are independently regulated in vitro and in human physiology by such factors as phosphorylation and the allosteric effect of cytochrome b5. P450c21 is a related steroidogenic enzyme that catalyzes the 21-hydroxylase reaction. P450c17 and P450c21 have high sequence similarity and identical gene structures, and both utilize the same substrates (pregnenolone, progesterone, and their 17-hydroxy derivatives). The subtle differences in their substrate selectivities and catalytic specificities lead to the production of different steroid hormones with vastly different biological activities. We believe that structure-function comparisons of the two enzymes will yield important insight to the actions of steroidogenic enzymes and approaches to their selective inhibition. We have completed a computer-graphic model of human P450c17 based on the X-ray structure of the bacterial P450BM-P using the resources at the UCSF Computer Graphics Laboratory. Based on our analysis of the P450c17 model using the CGL resources, we have designed chimeric proteins in an attempt to confer 21-hydroxylase activity to P450c17 and 17-hydroxylase activity to P450c21. We have also analyzed several structures of cytochrome b5 and apo-cytochrome b5 in the CGL. Based on this analysis, plus our biochemical studies, we are engineering mutant forms of b5 which we will use to probe the interaction of P450c17 with b5. We will use these data to refine the model and to dock protein surfaces together, with the goal of describing the structure of the entire catalytic complex.