Computational insights into the different catalytic activities of CYP2A13 and CYP2A6 on NNK

The human cytochrome P450 2A13 (CYP2A13) and P450 2A6 (CYP2A6) are 94% identical in amino acid sequence, but they metabolize many substrates with different efficiencies. Previous experimental results have shown that CYP2A13 exhibited catalytic activity that was more than 300-fold higher than CYP2A6 toward 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone (NNK), a carcinogen present in tobacco products. At present, however, the structural determinants accounting for the differential catalytic activities of these two isozymes toward NNK remain unclear. In the present study, molecular docking combined with molecular dynamics simulation and binding free energy calculation was performed to investigate the above issue. The results demonstrate that NNK was able to form a hydrogen bond with Asn297 in either CYP2A13 or CYP2A6. The hydrogen-bond acceptor was the pyridine nitrogen of NNK in the CYP2A13 complex, but it changed to the carbonyl oxygen in the CYP2A6 complex. NNK interacted with the residues in helix I and the K-β2 loop in CYP2A13, whereas it preferred to contact with the phenylalanine cluster in CYP2A6. The residues in helix I and the K-β2 loop of CYP2A13 played a vital role in keeping NNK in a more stable binding state. The binding free energies calculated by MM-GBSA were in agreement with the experimental results.