Exploration Of Special Metabolic Phenomenon In CYP450 Family

Cytochrome P450 protein superfamily are heme-containing monooxygenases, while the primary sequences of the family members vary, but they share common structure features. In particular, certain subtypes play key roles in human endogenous or exogenous substances metabolic process.In the CYP450 superfamily, with large active pocket size CYP3A4 often accommodates multiple substrates. The interaction between these substrates(i.e. ketoconazole) has certain effect on the metabolic process, showing special metabolic kinetics, but the mechanism has yet to be revealed.CYP2J2 is another important subtype, which plays an important role in fatty acid metabolism in human body. In the cardiovascular system, CYP2J2 can take arachidonic acid as the main substrate, generating epoxyeicosatrienoic acids(EETs). This product can protect endothelial cells from hypoxic or ischemic injury, closely associated with the pathogenesis of coronary artery disease and hypertension. Some mutants of CYP2J2(T143A and N404Y) significantly decrease the enzyme activity in catalytic reaction mentioned above, but the specific mechanism is not clear. Therefore, clarifying the mechanism is critical for treatment in coronary artery disease and hypertension.In this thesis, we applied molecular dynamics simulation method to explore the two major problems respectively. For the ketoconazole metabolism in CYP3A4, we found that pi-pi stacking between the two substrate molecules and protein stabilized the first substrate, thus accelerated the catalytic reaction. For CYP2J2, based on modeled structures of wild-type and two mutants, we found that T143 A mutation led to water channel closure, thus hindered the reaction process; in N404 Y mutation, the steric effect of the Tyr404 side-chain could reposition the side-chain of Leu378, making it no longer form a hydrogen bond with arachidonic acid, which would weaken the substrate binding affinity, so as to reduce the substrate specificity of CYP2J2. These results provide interpretation for the mechanism of the two issues above, but also lay the foundation for follow-up study.