Co-expression of human cytochrome P4501A1 (CYP1A1) variants and human NADPH-cytochrome P450 reductase in the baculovirus/insect cell system

Xenobiotica. 2001 Jun;31(6):345-56. doi: 10.1080/00498250110055947.


1. Three human cytochrome P4501A1 (CYP1A1) variants, wild-type (CYP1A1.1), CYP1A1.2 (1462V) and CYP1A1.4 (T461N), were co-expressed with human NADPH-P450 reductase (OR) in Spodoptera frugiperda (Sf9) insect cells by baculovirus co-infection to elaborate a suitable system for studying the role of CYPA1 polymorphism in the metabolism of exogenous and endogenous substrates. 2. A wide range of conditions was examined to optimize co-expression with regard to such parameters as relative multiplicity of infection (MOI), time of harvest, haem precursor supplementation and post-translational stabilization. tinder optimized conditions, almost identical expression levels and molar OR/CYP1A1 ratios (20:1) were attained for all CYP1A1 variants. 3. Microsomes isolated from co-infected cells demonstrated ethoxyresorufin deethlylase activities (nmol/min(-1) nmol(-1) CYP1A1) of 16.0 (CYP1A1.1), 20.5 (CYP1A1.2) and 22.5 (CYP1A1.4). Pentoxyresorufin was dealkylated approximately 10-20 times slower with all enzyme variants. 4. All three CYP1A1 variants were active in metabolizing the precarcinogen benzo[a]pyrene (B[a]P), with wild-type enzyme showing the highest activity, followed by CYP1A1.4 (60%) and CYP1A1.2 (40%). Each variant produced all major metabolites including B[a]P-7,8-dihydrodiol, the precursor of the ultimate carcinogenic species. 5. These studies demonstrate that the baculovirus-mediated co-expression-by-co-infection approach all CYP1A1 variants yields functionally active enzyme systems with similar molar OR/CYP1A1 ratios, thus providing suitable preconditions to examine the metabolism of and environmental chemicals by the different CY1A1 variants.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • Baculoviridae / genetics*
  • Benzo(a)pyrene / metabolism
  • Carcinogens / metabolism
  • Cytochrome P-450 CYP1A1 / genetics*
  • Cytochrome P-450 CYP1A1 / metabolism
  • Gene Expression*
  • Genetic Vectors
  • Humans
  • Kinetics
  • Microsomes / enzymology
  • Mutagenesis
  • NADPH-Ferrihemoprotein Reductase / genetics*
  • NADPH-Ferrihemoprotein Reductase / metabolism
  • Spodoptera / enzymology
  • Spodoptera / genetics*
  • Substrate Specificity
  • Transfection


  • Carcinogens
  • Benzo(a)pyrene
  • Cytochrome P-450 CYP1A1
  • NADPH-Ferrihemoprotein Reductase