Metabolism of benzo[a]pyrene to trans-7,8-dihydroxy-7, 8-dihydrobenzo[a]pyrene by recombinant human cytochrome P450 1B1 and purified liver epoxide hydrolase

Chem Res Toxicol. 1999 Jul;12(7):623-9. doi: 10.1021/tx990028s.


Recombinant human enzymes expressed in membranes obtained from Escherichia coli transformed with cytochrome P450 (P450) and NADPH-P450 reductase cDNAs were used to identify the human P450 enzymes that are most active in catalyzing the oxidative transformation of benzo[a]pyrene in vitro. Activation of benzo[a]pyrene to genotoxic products that cause induction of umu gene expression in Salmonella typhimurium NM2009 by P450 1A1 and P450 1B1 enzymes was found to be enhanced by inclusion of purified epoxide hydrolase (isolated from rat or human livers) with the reaction mixture. High-performance liquid chromatographic analysis showed that P450 1B1 catalyzed benzo[a]pyrene to trans-7, 8-dihydroxy-7,8-dihydrobenzo[a]pyrene at level of approximately 3 nmol min(-)(1) nmol of P450(-)(1) only when epoxide hydrolase was present and P450 1A1 (with the hydrolase) was able to catalyze benzo[a]pyrene at one-tenth of the activity catalyzed by P450 1B1. Kinetic analysis showed that ratio of V(max) to K(m) for the formation of trans-7,8-dihydroxy-7,8-dihydrobenzo[a]pyrene in this assay system was 3.2-fold higher in CYP1B1 than in CYP1A1. Other human P450s (including P450s 1A2, 2E1, and 3A4) were found to have very low or undetectable activities toward the formation of trans-7, 8-dihydroxy-7,8-dihydrobenzo[a]pyrene. A reconstituted system containing purified P450 1B1, rabbit liver NADPH-P450 reductase, and human liver epoxide hydrolase was found to catalyze benzo[a]pyrene to trans-7,8-dihydroxy-7,8-dihydrobenzo[a]pyrene at a rate of 0.86 nmol min(-)(1) nmol of P450(-)(1); the activities were found to be largely dependent on the presence of sodium cholate in the system. These results suggest that P450 1B1 is a principal enzyme in catalyzing the oxidation of benzo[a]pyrene to trans-7,8-dihydroxy-7, 8-dihydrobenzo[a]pyrene and that the catalytic functions of P450 1B1 may determine the susceptibilities of individuals to benzo[a]pyrene carcinogenesis.

Publication types

  • Research Support, Non-U.S. Gov't
  • Research Support, U.S. Gov't, P.H.S.

MeSH terms

  • Animals
  • Aryl Hydrocarbon Hydroxylases*
  • Benzo(a)pyrene / metabolism*
  • Benzo(a)pyrene / toxicity
  • Carcinogens / metabolism*
  • Carcinogens / toxicity
  • Chromatography, High Pressure Liquid
  • Cytochrome P-450 CYP1B1
  • Cytochrome P-450 Enzyme System / metabolism*
  • Dihydroxydihydrobenzopyrenes / metabolism*
  • Epoxide Hydrolases / metabolism*
  • Humans
  • Liver / enzymology*
  • Mutagenicity Tests
  • Rabbits
  • Rats
  • Recombinant Proteins / metabolism
  • Salmonella typhimurium / metabolism


  • Carcinogens
  • Dihydroxydihydrobenzopyrenes
  • Recombinant Proteins
  • benzo(a)pyrene 7,8-dihydrodiol
  • Benzo(a)pyrene
  • Cytochrome P-450 Enzyme System
  • Aryl Hydrocarbon Hydroxylases
  • CYP1B1 protein, human
  • Cyp1b1 protein, rat
  • Cytochrome P-450 CYP1B1
  • Epoxide Hydrolases