As a family of structurally-related enzymes, P450 exhibits extreme diversity in its activity and also in its gene regulation. We study the structural and molecular basis for the diverse activity and regulation, since the diversity affects the human susceptibility to naturally-occurring and man-made toxins and carcinogens. Using site-directed mutagenesis and heterologous expression system, we find that P450 activity can be altered as small as a single amino acid substitution; the substrate specificity of P4502A5 is converted from coumarin to testosterone and, then, to corticosterone by a single mutation of the residue-209 from Phe to Leu and then to Asn. In addition, the steroid-binding site and configuration in the hydrophobic heme-pocket in P450 is elucidated by incorporating the enzymatic characteristics into the computational graphics using the 3D of bacterial P450cam as the model. Transcriptional regulation by growth hormone (GH) is the principal mechanism of sex-specific expression of steroid hydroxylases P45015alpha and P45016alpha. Using the strain differences in the expression among the domestic and wild mice, we found a transacting locus which represses the transcription of female-specific P45015alpha gene in male mice. Transcriptional promoters are determined in the male-specific P45016alpha gene and sex-specific nuclear proteins binding to the promoters are identified. This progress in our research is providing the biological basis for understanding the P450 diversities and their effects on the differences in human susceptibility to environmental chemicals.
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