CYP2A6 uses a heme cofactor to oxidize the substrates. The active site of this enzyme is compact and composed by a hydrophobic Phe-cluster formed by residues Phe107, Phe111, Phe108,Phe209 and Phe480 (Fig. 1). In this region only one hydrogen bond donor is available and it is provided by Asn297, that directs the natural substrate (coumarin) towards regioselective oxidation [33].