Microsomal cytochrome P450 family 1 enzymes play prominent roles in xenobiotic detoxication and procarcinogen activation. P450 1A2 is the principal cytochrome P450 family 1 enzyme expressed in human liver and participates extensively in drug oxidations. This enzyme is also of great importance in the bioactivation of mutagens, including the N-hydroxylation of arylamines. P450-catalyzed reactions involve a wide range of substrates, and this versatility is reflected in a structural diversity evident in the active sites of available P450 structures. Here, we present the structure of human P450 1A2 in complex with the inhibitor ␣-naphthoflavone, determined to a resolution of 1.95 Å . ␣-Naphthoflavone is bound in the active site above the distal surface of the heme prosthetic group. The structure reveals a compact, closed active site cavity that is highly adapted for the positioning and oxidation of relatively large, planar substrates. This unique topology is clearly distinct from known active site architectures of P450 family 2 and 3 enzymes and demonstrates how P450 family 1 enzymes have evolved to catalyze efficiently polycyclic aromatic hydrocarbon oxidation. This report provides the first structure of a microsomal P450 from family 1 and offers a template to study further structure-function relationships of alternative substrates and other cytochrome P450 family 1 members. Enzymes of the cytochrome P450 (CYP)5 superfamily play a significant physiologic role in the detoxication of foreign compounds and the biosynthesis of endogenous compounds, including steroid hormones, bile acids, and cholesterol. The enzymes comprising P450 families 1, 2, and 3 contribute most extensively to the biotransformation of xenobiotics to more polar metabolites that are more readily excreted. In humans and most mammals, family 1 contains three well characterized P450 monooxygenases; 1A1, 1A2, and 1B1. These enzymes are generally distinguished from P450s in other families by their capacity to oxidize a variety of polynuclear aromatic hydrocarbons (PAHs).6 Moreover, the expression levels of the three enzymes are induced by exposure to PAHs (1). The induction is mediated by a ligand-activated transcription factor, the aryl hydrocarbon receptor, which is a basic-loop-helix PAS domain protein that binds to enhancer elements flanking the CYP1A1, CYP1A2, and CYP1B1 genes and stimulates transcription.The oxidation of PAHs is generally protective. However, some P450-catalyzed reactions can transform these relatively inert compounds into genotoxic metabolites that can initiate mutagenesis and cancer. Human P450 1A2 is notable among family 1 enzymes for the capacity to N-oxidize arylamines, the major metabolic process in the bioactivation of arylamines to potent mutagenic or carcinogenic compounds (2). ␣-Naphthoflavone (ANF), a prototype flavonoid, is known to competitively inhibit P450s of family 1, albeit at different concentrations, and has been used to discriminate between P450 family 1 enzymes (3). Flavonoids have gained recent interest in vi...

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Background: Knowledge of the structural features of P450 2C19 that underlie its distinct roles in human drug metabolism is lacking. Results: The structure of P450 2C19 was determined by x-ray crystallography. Conclusion:The structure of the enzyme exhibits features that distinguish it from closely related P450s 2C8 and 2C9. Significance: Structural characterization of P450 2C19 contributes to a better understanding of its role in drug clearance.

Our objective was to determine the structure of 2C19 and to identify structural features underlying the distinct substrate and inhibitor profiles of 2C19 and 2C9, which differ at 49 of 490 residues. Crystal structures of 2C19 and 2C9 complexed with structurally related inhibitors, DMB and DMI, respectively, were determined to 2.9 and 1.9 Angstrom resolution. DMI is the 3,5‐diiodophenyl analog of DMB (2‐methyl‐1‐benzofuran‐3‐yl)‐(4‐hydroxy‐3,5‐dimethylphenyl)methanone. The increased bulk of the iodine relative to the methyl moieties and the lower pKa of the phenol are thought to contribute to 1000‐fold difference in the Ki's exhibited by DMI for the two enzymes (Locuson et al. J. Med. Chem. 47: 6768 (2004)). The two enzymes exhibit conformation differences that confer substrate selectivity with relatively few amino acid differences contributing directly to substrate binding. The increased flexibility of the helix B‐C loop of 2C9 relative to 2C19 allows Arg108 to complement the negative charge of the phenol, whereas the neutral form of DMB is more readily accommodated by 2C19. These structures elucidate the contributions of structural flexibility and active site hydration in substrate binding. Furthermore, the availability of a structure for 2C19 will facilitate computational approaches for predictions of substrate and inhibitor binding this enzyme. Supported by USPHS grant NIH GM031001.

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