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Carcinogenesis, Vol. 20, No. 8, 1607-1614, August 1999
© 1999 Oxford University Press


Carcinogenesis

Catalytic properties of polymorphic human cytochrome P450 1B1 variants

Tsutomu Shimada5, Junko Watanabe1, Kaname Kawajiri1, Thomas R. Sutter2, F. Peter Guengerich3, Elizabeth M.J. Gillam4,5 and Kiyoshi Inoue

Osaka Prefectural Institute of Public Health, 3-69 Nakamichi 1-chome, Higashinari-ku, Osaka 537-0025 and
1 Saitama Cancer Center Research Institute, Kitaadachi-gun, Saitama 362-0806, Japan,
2 Division of Toxicological Sciences, Johns Hopkins University, School of Hygiene and Public Health, Baltimore, MD, 21205 and
3 Department of Biochemistry and Center in Molecular Toxicology, Vanderbilt University School of Medicine, Nashville, TN 37232, USA and
4 Department of Physiology and Pharmacology, The University of Queensland, St Lucia, Queensland 4072, Australia

Four polymorphic human cytochrome P450 (CYP) 1B1 allelic variants, namely Arg48,Ala119,Leu432,Asn453, Arg48,Ser119,Leu432,Asn453, Arg48,Ala119,Val432,Asn-453 and Arg48,Ser119,Val432,Asn453, were expressed in Escherichia coli together with human NADPH-P450 reductase and the recombinant proteins (in bacterial membranes) were used to assess whether CYP1B1 polymorphisms affect catalytic activities towards a variety of P450 substrates, including diverse procarcinogens and steroid hormones. Activities for activation of 19 procarcinogens to DNA-damaging products by these four CYP1B1 variants in a Salmonella typhimurium NM2009 umu response system were found to be essentially similar, except that a Arg48, Ser119,Leu432,Asn453 variant was slightly more active (1.2- to 1.5-fold) than the other three CYP1B1 enzymes in catalyzing activation of (+)- and (–)-benzo[a]pyrene-7,8-diols, 7,12-dimethylbenz[a]anthracene-3,4-diol, benzo[g]chrysene-11,12-diol, benzo[b]fluoranthene-9,10-diol,2-amino-3,5-dimethylimidazo[4,5-f]quinoline, 2-amino-3-methylimidazo[4,5-f]quinoline and 2-aminofluorene. Kinetic analysis of 17ß-estradiol hydroxylation showed that Vmax values for 4-hydroxylation ranged between 0.9 and 1.5 nmol/min/nmol P450 for 4-hydroxylation and 0.3 and 0.6 nmol/min/nmol P450 for 2-hydroxylation in these CYP1B1 variants, with Km values ranging from 1 to 9 µM. Interestingly, the ratio of product formation of 4-hydroxyestradiol to 2-hydroxyestradiol was higher for the Val432 variants of CYP1B1 variants than the Leu432 variants of the enzyme. The same trend was noted in the ratio of estrone 4-hydroxylation to estrone 2-hydroxylation catalyzed by CYP1B1 variants. Mutation in the CYP1B1 genes also affected the Km and Vmax values in the 6ß-hydroxylation of testosterone and 6ß- and 16{alpha}-hydroxylation of progesterone. These results indicate that the polymorphisms in the human CYP1B1 gene cause some alterations in catalytic function towards procarcinogens and steroid hormones and thus may make some contribution to susceptibilities of individuals towards mammary and lung cancers in humans.

Abbreviations: B[g]C, benzo[g]chrysene; B[b]F, benzo[b]fluoranthene; B[a]P, benzo[a]pyrene; B[c]P, benzo[c]phenanthrene; CYP, cytochrome P450; DB[a,l]P, dibenzo[a,l]pyrene; DMBA, 7,12-dimethylbenz[a]anthracene; hNPR, human NADPH-P450 reductase; IQ, 2-amino-3-methylimidazo [4,5-f]quinoline; MeIQ, 2-amino-3,5-dimethylimidazo[4,5-f]quinoline; MeIQx, 2-amino-3,8-dimethylimidazo[4,5-f]quinoxaline; Trp-P-1, 3-amino-1,4-dimethyl5H-pyrido[4,3-b]indole; diol, used in the text to designate the prefix `dihydroxydihydro' for individual polycyclic hydrocarbons.

5 To whom correspondence should be addressed Email: shimada{at}iph.pref.osaka.jp (T.S.); gillam{at}plpk.uq.edu.au (E.M.J.G.)


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Cytochrome P450 1B1 (CYP1B1) Pharmacogenetics: Association of Polymorphisms with Functional Differences in Estrogen Hydroxylation Activity
Cancer Res., July 1, 2000; 60(13): 3440 - 3444.
[Abstract] [Full Text]


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Cancer Epidemiol. Biomarkers Prev.Home page
W. Zheng, D.-W. Xie, F. Jin, J.-R. Cheng, Q. Dai, W.-Q. Wen, X.-O. Shu, and Y.-T. Gao
Genetic Polymorphism of Cytochrome P450-1B1 and Risk of Breast Cancer
Cancer Epidemiol. Biomarkers Prev., February 1, 2000; 9(2): 147 - 150.
[Abstract] [Full Text]



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