Hcooh

Balancing Step 3

FIGURE 2-30 Proposed mechanism for the aromatization reaction mediated by P450aro. The reactions require an appropriate A ring as substrate (either testosterone or androst-4-ene-3,17-dione), 3 mol of NADPH, and three molecules of 02. The accumulated evidence indicates that the initial hydroxylation occurs on carbon-19, followed immediately by a second hydroxylation on carbon-19 to yield a gem-diol, which fragments into a C-19 aldehyde. The third oxidative reaction is proposed to involve a peroxidative attack on the C-19 carboxyl group, as indicated previously. Of the three oxygen insertions, only the third is shown in detail. The model then proposes the participation of a COCT from glutamate and an NH2+ from lysine or histidine of the aromatase to facilitate 2/3-hydrogen elimination, followed by 1/3-hydrogen elimination and enolization of the 3-ketone, resulting in the production of an aromatic A ring. Adapted with permission from Simpson, E. R. et al. (1994). Aromatase cytochrome P-450, the enzyme responsible for estrogen biosynthesis. Endocr. Rev. 15, 342-355.

dence has accumulated to support the existence of specific regulatory elements in the promoter regions of the genes of these steroid-metabolizing enzymes. For example, the promoter of P450cll coding has been found to contain multiple czs-acting elements, termed Adl, Ad2, Ad3, Ad4, Ad5, and Ad6. The Ad4BP motif has also been found in the promoter of other steroid hydroxylases. The Ad4-binding protein (Ad4BP) has been cloned and sequenced; it codes for a protein of 51,000 that has a zinc finger domain analogous to the zinc finger of steroid receptors and belongs to the steroid receptor superfamily (see Figure 1-42). It seems likely that Ad4BP plays an important role in the regulation of gene expression of selective P450 enzymes.

10. Cytochrome P450 Gene Families

To date there are at least 221 known P450 genes that have been described in 31 eukaryotes (including 11 mammalian and 3 plant species) and 11 prokaryotes. All of these proteins are encoded by a superfamily of P450 genes that have their origins more than 3.5 billion years ago. This superfamily comprises at least 10 separate gene families; a diagram of the evolutionary relationships of the P450 genes is provided in Figure 231. This figure illustrates that despite the similarity of biological roles and steroidal substrates of the four adrenal steroidogenic enzymes located in the endoplasmic reticulum, P450aro, P450cl7, P450c21A, and P450c21B belong to four different gene families that are only quite distantly related to the two mitochondrial P450 enzymes, P450scc and P450cll. It appears that the highly coordinated pathways of steroid metabolism (see Figure 2-18) represent the convergence of enzymatic functions that arose at grossly different times in evolution.

C. Other Enzymes of Steroid Metabolism

1. 3ß-Hydroxy Dehydrogenase/A5,bt-Isomerase

A key enzymatic transformation in the biosynthetic pathway of five steroid hormones is the conversion of the 5-ene 3/3-hydroxysteroid to a 4-ene 3-oxosteroid

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