Characterization of cytochrome P-450 2B6 in human liver microsomes.
A cytochrome P-450 (P-450) enzyme of the CYP2B subfamily was partially purified from human liver microsomes and characterized with respect to immunochemical properties, N-terminal amino acid sequence, and catalytic activities toward typical P-450 substrates. P-450 enzymes were monitored in chromatographic fractions by immunoblotting analysis using antibodies raised against a monkey P-450 2B, as well as several purified human P-450 enzymes. The final P-450 2B preparation thus obtained was contaminated with P-450 3A4, but an N-terminal amino acid sequence matching the sequence predicted from the CYP2B6 cDNA was obtained. The apparent M(r) of this protein was 48 kDa, and the migration on sodium dodecyl sulfate-polyacrylamide gel electrophoresis was the same as that of the P-450 2B6 protein expressed in a human lymphoblast cell line. Immunoblotting analysis of 50 human liver samples revealed that the protein band considered to be P-450 2B6 was detected in only 12 samples, with four of these having relatively high levels. Several activities toward typical P-450 substrates were determined in a reconstituted monooxygenase system containing partially purified P-450 2B6 and compared with those obtained using a highly purified preparation of P-450 3A4 enzyme; we found that most of the activities were similar in these preparations, except that the partially purified P-450 2B6 showed high rates of activation of the mutagens 6-aminochrysene and 3-methoxy-4-aminoazobenzene to genotoxic metabolites in Salmonella typhimurium NM2009 strain.(ABSTRACT TRUNCATED AT 250 WORDS)
Mimura
M
M
Osaka Prefectural Institute of Public Health, Japan.
Baba
T
T
Yamazaki
H
H
Ohmori
S
S
Inui
Y
Y
Gonzalez
F J
FJ
Guengerich
F P
FP
Shimada
T
T
D000595:Amino Acid Sequence; D000970:Antineoplastic Agents / Q000493:pharmacokinetics; D001711:Biotransformation; D002273:Carcinogens / Q000493:pharmacokinetics; D002911:Chrysenes / Q000493:pharmacokinetics; D003577:Cytochrome P-450 Enzyme System / Q000378:metabolism*; D006801:Humans; D007150:Immunohistochemistry; D007527:Isoenzymes / Q000378:metabolism*; D008214:Lymphocytes / Q000201:enzymology / Q000378:metabolism; D008862:Microsomes, Liver / Q000201:enzymology*; D008969:Molecular Sequence Data; D010128:p-Aminoazobenzene / Q000031:analogs & derivatives / Q000493:pharmacokinetics
D000970:Antineoplastic Agents; D002273:Carcinogens; D002911:Chrysenes; D007527:Isoenzymes; D010128:p-Aminoazobenzene; C044436:3-methoxy-4-aminoazobenzene; D003577:Cytochrome P-450 Enzyme System; C100101:6-chrysenamine