Purification of a human cytochrome P-450 isozyme catalyzing lanosterol 14 alpha-demethylation.
An isozyme of cytochrome P-450 catalyzing lanosterol 14 alpha-demethylation was purified from human liver using column chromatography, including immunoaffinity chromatography. The purified protein exhibited a single protein band (53 kDa) on sodium dodecylsulfate-polyacrylamide gel electrophoresis. When reconstituted with NADPH-cytochrome P-450 reductase, the purified protein showed an activity of 14 alpha-demethylation of 24,25-dihydrolanosterol (3.20 nmol/min per mg protein). The apparent Km value for 24,25-dihydrolanosterol was found to be 27 microM. This enzyme converted in the reconstituted system, the oxygenated intermediates of 24,25-dihydrolanosterol 14 alpha-demethylation, 32-hydroxy-24,25-dihydrolanosterol and 32-oxo-24,25-dihydrolanosterol, to the 32-nor compound, 4,4-dimethylcholesta-8,14-dien-3 beta-ol.
Kyoritsu College of Pharmacy, Tokyo, Japan.
Y
Y
Sonoda
M
M
Endo
K
K
Ishida
Y
Y
Sato
N
N
Fukusen
M
M
Fukuhara
D000818:Animals; D000906:Antibodies / Q000276:immunology; D003429:Cross Reactions; D003577:Cytochrome P-450 Enzyme System / Q000276:immunology / Q000302:isolation & purification* / Q000378:metabolism; D006801:Humans; D007810:Lanosterol / Q000031:analogs & derivatives / Q000378:metabolism; D008862:Microsomes, Liver / Q000201:enzymology*; D010088:Oxidoreductases / Q000276:immunology / Q000302:isolation & purification* / Q000378:metabolism; D058886:Sterol 14-Demethylase; D013552:Swine
D000906:Antibodies; C548418:CYP51A1 protein, human; D007810:Lanosterol; C015316:lanostenol; D003577:Cytochrome P-450 Enzyme System; D010088:Oxidoreductases; D058886:Sterol 14-Demethylase