[Heterologous expression of functionally active human cytochrome p-450s. II. Modeling the metabolism of xenobiotics using Saccharomyces cerevisiae cells, expressing cytochrome P-450IIIA4].
cDNA encoding human cytochrome P-450IIIA4 was modified by the PCR method to attain maximum expression in the Saccharomyces cerevisiae cells and then recloned in the shuttle vector pYeDP under the control of galactose-sensitive promoter. The strain transformed this plasmid synthesized in the galactose-containing medium a protein authentic to P-450IIA4 in its spectral properties and enzymatic activity. After subcellular fractionation of the yeast the protein was found in the microsomal fraction which displayed a reduced-CO spectrum with maximum absorption at 448 nm typical of all members of the P-450 family. The microsomes from the transformed yeast catalyzed transformation of nifedipine, a specific substrate for P-450IIA4. Thus, transgenic yeast have been constructed mimicking the catabolic activity of human cytochrome P-450IIIA4.
Krynetskiĭ
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D001483:Base Sequence; D003001:Cloning, Molecular; D051544:Cytochrome P-450 CYP3A; D003577:Cytochrome P-450 Enzyme System / Q000235:genetics / Q000378:metabolism*; D018076:DNA, Complementary; D005822:Genetic Vectors; D006801:Humans; D006899:Mixed Function Oxygenases / Q000235:genetics / Q000378:metabolism*; D008954:Models, Biological; D008969:Molecular Sequence Data; D011401:Promoter Regions, Genetic; D012441:Saccharomyces cerevisiae / Q000235:genetics*; D015262:Xenobiotics / Q000378:metabolism*
D018076:DNA, Complementary; D015262:Xenobiotics; D003577:Cytochrome P-450 Enzyme System; D006899:Mixed Function Oxygenases; C104464:CYP3A protein, human; D051544:Cytochrome P-450 CYP3A
Geterologicheskaia ékspressiia funktsional'no aktivnykh tsitokhromov P-450 cheloveka. II. Modelirovanie metabolizma ksenobiotikov s pomoschch'iu kletok Saccharomyces cerevisia, ékspressiruiushchikh tsitokhrom P-450IIIA4.