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Arginine to lysine 108 substitution in recombinant CYP1A2 abolishes methoxyresorufin metabolism in lymphoblastoid cells
Author(s) -
Hadjokas Nicholas E,
Dai Renke,
Friedman Fred K,
Spence Michael J,
Cusack Barry J,
Vestal Robert E,
Ma Yongsheng
Publication year - 2002
Publication title -
british journal of pharmacology
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 2.432
H-Index - 211
eISSN - 1476-5381
pISSN - 0007-1188
DOI - 10.1038/sj.bjp.0704711
Subject(s) - cyp1a2 , biochemistry , chemistry , mutagenesis , active site , binding site , enzyme , stereochemistry , cytochrome p450 , mutation , gene
Cytochrome P4501A2 (CYP1A2) activates a large number of procarcinogens to carcinogens. Phytochemicals such as flavones can inhibit CYP1A2 activity competitively, and hydroxylated derivatives of flavone (galangin) may be potent, selective inhibitors of CYP1A2 activity relative to CYP1A1 activity. Molecular modelling of the CYP1A2 interaction with hydroxylated derivatives of flavone suggests that a number of hydrophobic residues of the substrate‐binding domain engage in hydrogen bonding with such inhibitors. We have tested this model using site‐directed mutagenesis of these residues in expression plasmids transfected into the human B‐lymphoblastoid cell line, AHH‐1 TK+/−. Consistent with the molecular model's predicted placement in the active site, amino acid substitutions at the predicted residues abolished CYP1A2 enzymatic activity. Transfected cell lines contained equal amounts of immunoreactive CYP1A2. Our results support the molecular model's prediction of the critical amino acid residues present in the hydrophobic active site, residues that can hydrogen bond with CYP1A2 inhibitors and modify substrate binding and/or turnover.British Journal of Pharmacology (2002) 136 , 347–352; doi: 10.1038/sj.bjp.0704711