Structural insight into the altered substrate specificity of human cytochrome P450 2A6 mutants

Arch Biochem Biophys. 2007 Aug 15;464(2):197-206. doi: 10.1016/ Epub 2007 May 11.


Human P450 2A6 displays a small active site that is well adapted for the oxidation of small planar substrates. Mutagenesis of CYP2A6 resulted in an increased catalytic efficiency for indole biotransformation to pigments and conferred a capacity to oxidize substituted indoles (Wu, Z.-L., Podust, L.M., Guengerich, F.P. J. Biol. Chem. 49 (2005) 41090-41100.). Here, we describe the structural basis that underlies the altered metabolic profile of three mutant enzymes, P450 2A6 N297Q, L240C/N297Q and N297Q/I300V. The Asn297 substitution abolishes a potential hydrogen bonding interaction with substrates in the active site, and replaces a structural water molecule between the helix B'-C region and helix I while maintaining structural hydrogen bonding interactions. The structures of the P450 2A6 N297Q/L240C and N297Q/I300V mutants provide clues as to how the protein can adapt to fit the larger substituted indoles in the active site, and enable a comparison with other P450 family 2 enzymes for which the residue at the equivalent position was seen to function in isozyme specificity, structural integrity and protein flexibility.

Publication types

  • Research Support, N.I.H., Extramural

MeSH terms

  • Amino Acid Substitution
  • Aryl Hydrocarbon Hydroxylases / chemistry*
  • Aryl Hydrocarbon Hydroxylases / genetics
  • Aryl Hydrocarbon Hydroxylases / ultrastructure*
  • Binding Sites
  • Computer Simulation
  • Cytochrome P-450 CYP2A6
  • Enzyme Activation
  • Humans
  • Mixed Function Oxygenases / chemistry*
  • Mixed Function Oxygenases / genetics
  • Mixed Function Oxygenases / ultrastructure*
  • Models, Chemical*
  • Models, Molecular*
  • Mutagenesis, Site-Directed
  • Protein Binding
  • Protein Conformation
  • Structure-Activity Relationship
  • Substrate Specificity


  • Mixed Function Oxygenases
  • Aryl Hydrocarbon Hydroxylases
  • CYP2A6 protein, human
  • Cytochrome P-450 CYP2A6

Associated data

  • PDB/I300V
  • PDB/L240C
  • PDB/N297Q