Characterization of human lung microsomal cytochrome P-450 1A1 and its role in the oxidation of chemical carcinogens.
Rat and human lung microsomal cytochrome P-450 (P-450) enzymes have been characterized with regard to their catalytic activities towards several xenobiotic chemicals, including procarcinogens, in different microsomal preparations. Rat lung microsomal P-450s were more active than the human P-450s in catalyzing most of the monooxygenation reactions. Human lung microsomal P-450 was solubilized and purified. Human lung microsomes contain approximately 10 pmol of P-450/mg of protein, on the basis of Fe2+.CO versus Fe2+ difference spectra of the eluates obtained from an octylamino-agarose column. The partially purified P-450 preparations from two human lung microsomal samples showed high activities for the conversion of both (+)- and (-)-isomers of 7,8-dihydroxy-7,8-dihydrobenzo(a)pyrene to genotoxic products. After DEAE-cellulose column chromatography, a partially purified P-450 fraction containing polypeptides of Mr 52,000 and 58,000 was obtained from the early fraction of the octylamino-agarose column eluate, and an electrophoretically homogeneous protein having a molecular weight of approximately 52,000 was recovered from a latter fraction. The amino-terminal amino acid sequences of the two peptides in the earlier fraction were determined; neither polypeptide appears to resemble any known P-450 protein. The protein from the latter octylamino-agarose fraction was immunoreactive with anti-rat P-450 1A2 and anti-human P-450 1A2 but not with antibodies raised against other P-450 enzymes or autoimmune antibodies that specifically recognize human P-450 1A2. A tryptic peptide was isolated from the preparation, and the amino acid sequence matched that of human P-450 1A1 perfectly (residues 31-48) but not that of human P-450 1A2. All of nine human lung microsomal samples examined contained proteins that were immunoreactive with rabbit anti-rat P-450 1A2 and catalyzed the activation of 7,8-dihydroxy-7,8-dihydrobenzo(a)pyrene. The activities could be inhibited by rabbit anti-rat P-450 1A2 and, to a lesser extent, by anti-rat P-450 1A1. The addition of 7,8-benzoflavone caused inhibition or stimulation, depending upon the particular human lung microsomal preparation. Thus, this work clearly shows that human lung microsomes contain at least two major P-450 enzymes; human P-450 1A1 is present in lungs and can actually catalyze the activation of environmental procarcinogens, including polycyclic aromatic hydrocarbons.
Shimada
T
T
Department of Biochemistry, Vanderbilt University School of Medicine, Nashville, Tennessee 37232-0146.
Yun
C H
CH
Yamazaki
H
H
Gautier
J C
JC
Beaune
P H
PH
Guengerich
F P
FP
D000818:Animals; D002273:Carcinogens / Q000378:metabolism*; D002848:Chromatography, DEAE-Cellulose; D019388:Cytochrome P-450 CYP1A2; D003577:Cytochrome P-450 Enzyme System / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D004591:Electrophoresis, Polyacrylamide Gel; D006801:Humans; D008168:Lung / Q000201:enzymology*; D008861:Microsomes / Q000201:enzymology*; D008862:Microsomes, Liver / Q000201:enzymology; D010084:Oxidation-Reduction; D010088:Oxidoreductases / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D051381:Rats; D011994:Recombinant Proteins / Q000302:isolation & purification / Q000378:metabolism; D013379:Substrate Specificity
D002273:Carcinogens; D011994:Recombinant Proteins; D003577:Cytochrome P-450 Enzyme System; D010088:Oxidoreductases; D019388:Cytochrome P-450 CYP1A2