title: Effect of histidine-tag and R457H and E580Q mutations on catalytic activity of recombinant human cytochrome P450 oxidoreductase.
abstract: Cytochrome P450 oxidoreductase (POR) transfers electrons from NADPH to several oxygenase enzymes including cytochrome P450 (CYP). Genetic mutations in the POR gene have recently been identified and associated with an autosomal recessive genetic disease. In this study, Vmax, Km, and Vmax/Km values of cytochrome c reduction and NADPH oxidation activities for R457H variant, histidine-tagged wild-type, and histidine-tagged E580Q were compared with those for wild-type. Vmax/Km values of cytochrome c reduction for the R457H variant and histidine-tagged wild-type were 8% and 26%, respectively, of wild-type, whereas Vmax/Km values of NADPH oxidation for the R457H variant and histidine-tagged wild-type were similar to those for wild-type. The kinetic parameters of the histidine-tagged E580Q variant were similar to those for histidine-tagged wild-type, suggesting that E580Q mutation may be of minor importance in interindividual variation in drug response. These results suggest that R457H but not E580Q is essential for the deficiency of POR activities and that the histidine-tagged system would be inappropriate for POR function.
authors: Division of Natural Science, Osaka Kyoiku University, Kashiwara, Japan.
authors: Toshiro
authors: T
authors: Niwa
authors: Katsuhito
authors: K
authors: Minami
authors: Masanao
authors: M
authors: Katagiri
mesh_terms: D019943:Amino Acid Substitution; D045304:Cytochromes c / Q000378:metabolism; D006639:Histidine / Q000235:genetics / Q000378:metabolism*; D006801:Humans; D007700:Kinetics; D016297:Mutagenesis, Site-Directed; D050505:Mutant Proteins / Q000378:metabolism*; D009249:NADP / Q000378:metabolism; D009251:NADPH-Ferrihemoprotein Reductase / Q000235:genetics* / Q000378:metabolism*; D010084:Oxidation-Reduction; D010597:Pharmacogenetics / Q000379:methods; D011994:Recombinant Proteins / Q000378:metabolism
chemical_list: D050505:Mutant Proteins; D011994:Recombinant Proteins; C033223:polyhistidine; D006639:Histidine; D009249:NADP; D045304:Cytochromes c; D009251:NADPH-Ferrihemoprotein Reductase