CYP reductase

Figure 9.1 CYP catalytic cycle. The sequential two-electron reduction of CYP and the various transient intermediates were first described in the late 1960s [206], The sequence of events that make up the CYP catalytic cycle is shown. The simplified CYP cycle begins with heme iron in the ferric state. In step (i), the substrate (R—H) binds to the enzyme, somewhere nearthe distal region of the heme group and disrupts the water lattice within the enzymes active site [207], The loss of water elicits a change in the heme iron spin state (from low spin to high spin) [208]. Step (ii) involves the transfers of an electron from NADPH via the accessory flavoprotein NADPH-CYP reductase, with the electron flow going from the reductase prosthetic group FAD to FMN to the CYP enzyme [206,209]. The...

Cytochrome b5 affects the kinetics of drug metabolism by certain CYP enzymes hence, coexpression of this microsomal hemoprotein (together with NADPH-CYP reductase) can affect the catalytic efficiency of certain recombinant CYP enzymes (76,109). For example, the presence of cytochrome b5 tends to increase Fmax for reactions catalyzed by CYP3 A4, whereas it tends to decrease Km for reactions catalyzed by CYP2E1. In both cases, cytochrome b5 increases Vmax/Km, which is a measure of in vitro intrinsic clearance. The fact that some commercially available recombinant CYP enzymes are expressed with cytochrome b5 while others are not complicates the interpretation of results of studies performed with recombinant human CYP enzymes. 

Contains all liver microsomal CYP enzymes with physiological levels of cytochrome b5 and NADPH-CYP reductase. Establishes the degree of interindividual variation in metabolic formation or substrate disappearance. 

The absence of cytochrome bs and/or artificially high levels of NADPH-CYP reductase can alter the kinetics of CYP enzymes and/or introduce artifacts. 

Chloramphenicol and secobarbital exhibit properties similar to those of tienilic acid, but they have not been studied in humans (11). Oxidative dechlorination of chloramphenicol with formation of reactive acyl chlorides appears to be an important metabolic pathway for irreversible inhibition of CYP. Chloramphenicol binds to CYP, and subsequent substrate hydroxylation and product release are not impaired. The inhibition of CYP oxidation and the inhibition of endogenous NADPH oxidase activity suggest that some modification of the CYP has taken place, which inhibits its ability to accept electrons from the CYP reductase (11). Secobarbital completely inactivates rat CYP2B1 functionally, with partial loss of the heme chromophore. Isolation of the N-alkylated secobarbital heme adduct and the modified CYP2B1 protein revealed that the metabolite partitioned between heme N-alkylation, CYP2B1 protein modification, and epoxidation. A small fraction of the prosthetic heme modifies the protein and contributes to the CYP2B1 inactivation (12). 

Endothelial biotransformation (e.g, gamma-GTP, GST, NADPH-cyp reductase, MAO, UGT, alkaline phosphatase, epoxide hydrolases, AAD, COMT, BChE, etc)... 

Tirapazamine is inactivated by two-electron reduction steps catalyzed by quinone reductase, yielding first the mono-N-oxide (reaction a and compound 18). In contrast, it is activated to a cytotoxic nitroxide (16) by a one-electron reduction catalyzed by NADPH-cytochrome P450 reductase (reaction b). This delocalized radical loses one molecule of water to yield a reactive radical (reaction c and compound 17). Radical 17 can then abstract one hydrogen radical from DNA (reaction d and compound 18), leading to DNA breaks and cytotoxicity. In summary, both inactivation and activation involve reduction reactions, but cytotoxicity will depend on the relative levels of quinone reductase and CYP reductase in hypoxic cells. 

Those include C-, N- and 5-oxidations, N-, 0- and 5-deaIkylation, deaminations, and certain dehalogenations. Under anaerobic conditions, it can also catalyze reductive reactions. The CYP monooxygenase system is a multien-zymatic complex constituted by the CYP hemoprotein, the flavoprotein enzyme NADPH CYP reductase, and the unsatnrated phospholipid phosphatidylcholine. The isoforms involved in xenobiotic metabolism are membrane bonnd enzymes situated in the endoplasmic reticnlnm. After... 

N. crassa -CYP55 (P450 J from Fusarium oxysporum represented a new class of CYP, as it is soluble, and carries out nitric oxide reduction without the need for a CYP-reductase (CPR) or a requirement for molecular oxygen. It was the first eukaryotic CYP to have its structure resolved and was a member of a new class of CYP. ... 

Individual drug metabolizing enzymes DMEs) like CYPs, FMOs, and UGTs are now routinely expressed, usually in insect cells using a baculovirus. Ultracentrifugation provides vesicles called microsomes, which contain these membrane-bound DMEs. One popular type of recombinant CYP preparation called Supersomes comes with CYP reductase, a necessary cofactor, co-expressed. Not only individual CYPs (2B6, 3A4, etc.) but also allelic variants like CYP2C9 2 are available for the study of possible pharmacogenomic effects on metabolism. 

Ionic reduction using a hydride occurs in vivo during the reduction catalyzed by NADH or NADPH enzymes, whereas one-electron reduction releases a radical structure, which may contribute to the toxic effect. Figure 33.13 illustrates the biotransformations affecting the anthracycline antitumor drug daunomycin. Recent smdies suggest that nitric oxide synthases may contribute to the cardiotoxicity, probably because of their structural similarities with CYP reductase. ... 

Xanthin oxidase (XO) Flavin-containing monooxygenase (FMO) Monoamine oxidase (MO) Aldehyde dehydrogenase CYP reductase Carbonyl reductase Epoxide hydrolase Esterases... 

The protein levels of normally expressed CYPs 1A2, 2B1/2, and 2E1 increased significantly in liver microsomes from rats treated with the pyrrolizi-dine alkaloid retrorsine (12,18-dihydroxysenecio-nan-ll,16-dione 3-longilobine 30 mg/kg i.p.) compared to untreated control rats (P <0.05), but protein levels of CYP 4A3, CYP 3A1, and CYP reductase were unchanged after retrorsine treatment (Gordon et al. 2000). In addition, CYP lAl mRNA and protein, which are not detectable in the livers of control rats, were induced after retrorsine exposure. 

Activation by NADPH CYP reductase leading to formation of DNA adducts mutagenic and carcinogenic DNA intercalation inhibition of DNA polymerase, reverse transcriptase... 

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