Identification of catalase in human livers as a factor that enhances phenytoin dihydroxy metabolite formation by human liver microsomes.
We have reported previously that the formation of a 3',4'-dihydroxylated metabolite of phenytoin (3',4'-diHPPH) by human liver microsomal cytochrome P450 (P450) is enhanced by the addition of human liver cytosol [Komatsu et al., Drug Metab Dispos 2000;28:1361-8]. The enhancing factor was determined in this study. The addition of cytosolic proteins precipitated by 50% ammonium sulfate to incubation mixtures increased the rate of microsomal 3',4'-diHPPH formation. This fraction was separated further by diethylaminoethyl-, carboxymethyl-, and hydroxyapatite-column chromatography. The amino acid sequence of the purified protein of approximately 55kDa by electrophoresis revealed this protein to be a catalase. The addition of purified or authentic catalase to the incubation mixtures increased the rates of microsomal 3',4'-diHPPH formation from 3'- and 4'-hydroxylated metabolites and from phenytoin in a concentration-dependent manner. In reconstituted systems containing CYP2C9, CYP2C19, and CYP3A4, the formation of 3',4'-diHPPH was also enhanced by catalase to different extents. This is the first report that catalase in livers enhances drug oxidation activities catalyzed by P450 in human liver microsomes.
Division of Drug Metabolism, Faculty of Pharmaceutical Sciences, Kanazawa University, Kanazawa, Japan.
Tomoko
T
Komatsu
Hiroshi
H
Yamazaki
Miki
M
Nakajima
Tsuyoshi
T
Yokoi
D002374:Catalase / Q000378:metabolism*; D003577:Cytochrome P-450 Enzyme System / Q000378:metabolism; D003600:Cytosol / Q000201:enzymology / Q000378:metabolism; D006801:Humans; D006900:Hydroxylation; D008099:Liver / Q000166:cytology / Q000201:enzymology*; D008862:Microsomes, Liver / Q000201:enzymology / Q000378:metabolism*; D010672:Phenytoin / Q000031:analogs & derivatives* / Q000378:metabolism*; D011994:Recombinant Proteins / Q000378:metabolism
D011994:Recombinant Proteins; C001419:hydroxyphenytoin; C027364:5-(3-hydroxyphenyl)-5-phenylhydantoin; C038540:5-(3,4-dihydroxyphenyl)-5-phenylhydantoin; D010672:Phenytoin; D003577:Cytochrome P-450 Enzyme System; D002374:Catalase