Cytochrome P450 BM3 (P450 102) from Bacillus megaterium is a unique bacterial P450, formed from the fusion of a fatty acid hydroxylase to a eukaryotic-like NADPH-cytochrome P450 reductase flavoprotein in a single (1 19 kDa) polypeptide chain (1, 2). It is an attractive model system for enzymological and structural studies due to its homology with mammalian drug metabolising P450 systems, and wi...

The P450 2A6 catalyzed 7-hydroxylation of coumarin proceeded with a mean Km of 0.40 (+/-0.13) microM and Vmax of 6.34 nmol/nmol P450/min (36-fold variation) in microsomal preparations from a panel of 12 human livers. Substrate depletion was avoided during the kinetic determinations. 8-Methoxypsoralen (8-MOP) is a potent mechanism-based inactivator of human liver P450 2A6 and reconstituted purif...

The human cytochrome P450 2J2 catalyzes an epoxygenase reaction to oxidize various fatty acids including arachidonic acid. In this study, three recombinant enzyme constructs of P450 2J2 were heterologously expressed in Escherichia coli and their P450 proteins were successfully purified using a Ni(2+)-NTA affinity column. Deletion of 34 amino acid residues in N-terminus of P450 2J2 enzyme (2J2-D...

In this paper we demonstrate that NADPH-initiated oxidative damage of microsomal proteins occurs in the absence of free metal ions and that this protein oxidation is mediated by cytochrome P450 (cyt P450). Oxidized proteins are rapidly degraded by proteases. Ascorbate (AH2) specifically inhibits free metal ion-independent cyt P450-mediated protein oxidation and thereby prevents subsequent prote...

Although a single binary functional complex between cytochrome P450 (P450 or CYP for a specific isoform) and cytochrome P450 reductase (CPR) has been generally accepted in the literature, this simple model failed to explain the experimentally observed catalytic activity of recombinant CYP2E1 in dependence on the total concentration of the added CPR-K56Q mutant. Our rejection of the simplest 1:1...

This report summarizes a symposium sponsored by the American Society for Pharmacology and Experimental Therapeutics at Experimental Biology held April 20-24 in Boston, MA. Presentations discussed the status of cytochrome P450 (P450) knowledge, emphasizing advances and challenges in relating structure with function and in applying this information to drug design. First, at least one structure of...

The metabolism of arylhydrazines by cytochromes P450 (P450s) has previously been shown to yield aryl-iron complexes that inhibit P450 enzymes as a result of heme modification. These modifications of the heme have been used to probe the topology of the active site of several P450s. Therefore, diaziridines containing one or more substitutions on the phenyl ring were synthesized and evaluated as p...

Many undesired side effects or therapeutic failures of drugs are the result of differences or changes in drug metabolism, primarily depending on the levels and activities of cytochrome P450 (P450) enzymes. To assess whether P450 expression profiles can reflect hepatic drug metabolism, we compared P450 mRNA levels in the liver or peripheral leukocytes with the corresponding hepatic P450 activiti...