| CYP3A4 and CYP3A5 are important members of the cytochrome P450 family.Their overall structure is very similar,but the local structure of the active site is different.The drugs metabolized by these two enzymes overlap,and in many drug metabolism,CYP3A5 has a higher catalytic efficiency.The expression and activity of CYP3A4 are not affected by genetic factors,while CYP3A5 is only expressed in about 20% of the population due to the influence of gene phenotype,resulting in significant individual differences in drug efficacy and toxicity.In drug development,attention is often paid to the selective metabolism of CYP3A4.Research on CYP3A5 mainly focuses on genetic polymorphism and pharmacokinetics,and the mechanisms that determine the different catalytic efficiency of CYP3A4 and CYP3A5 have not been fully elucidated.Existing experimental studies can to some extent reveal the reaction mechanism between proteases and small drug molecules,but the results obtained through experimental methods have certain limitations and are difficult to capture dynamic information of physiological changes.Molecular dynamics simulation can compensate for this limitation by revealing the dynamic conformational changes between proteases and small drug molecules,while also capturing various energy information during the conformational change process.This paper selects three small molecule drugs,midazolam,carbamazepine,and amoxicillin,as substrates to bind to two p450 subtypes of CYP3A4 and CYP3A5,respectively.From an atomic perspective,the differences in CYP3A4 and CYP3A5 binding to the same substrate and their structure-function relationships are interpreted.The results obtained provide theoretical support for the research and development of specific isomer inhibitors for individuals with different CYP3A5 expression in the era of precision medicine.The main research elements of this thesis are as follows:1.Molecular dynamics simulation of the interaction between midazolam with two CYP isoforms 3A4 and 3A5CYP3A4 and CYP3A5 are two important members of cytochrome P450 family.Although their overall structures are similar,the local structures of the active site are different,which directly leads to obvious individual differences in drug metabolic efficacy and toxicity.In this work,midazolam(MDZ)was selected as the labeled substrate,and its interaction with two proteins,CYP3A4 and CYP3A5,was studied by molecular dynamics simulation(MD)along with the calculation of the binding free energy.The results show that CYP3A4 and CYP3A5 have similar enzyme-substrate binding patterns.In both complexes,Ser119 forms a high occupancy hydrogen bond with MDZ,which plays a key role in the stability of the interaction between MDZ and the enzymes.In addition,the complex formed by CYP3A4 and MDZ is more stable,which may be attributed to the sandwich structure formed by the fluorophenyl group of the substrate with Leu216 and Leu482.Our study interprets the binding differences between two isoform-substrate complexes and reveals structure function-relationship from the atomic perspective,which is expected to provide a theoretical basis for accurately measuring the effectiveness and toxicity of drugs for individuals in the era of precision medicine.2.Molecular dynamics simulation of the interaction between carbamazepine with two CYP isoforms 3A4 and 3A5Carbamazepine(CBZ),a commonly prescribed antiepileptic drug,is mainly metabolized by two isoforms of cytochrome P450(CYP),CYP3A4 and CYP3A5,in human liver.Therefore,the binding of CBZ with two enzymes plays crucial role in the biotransformation of the drug into its active metabolite.In the present work,molecular dynamics(MD)simulation was used to investigate the detailed interacction between CBZ and two CYP isoforms at the atomic level.The results reveal that although CBZ can bind with the two proteins,all kinds of the interactions,including hydrogen bonds,salt bridges,hydrophobic interaction,and π-π interactions,are isoform specific.It directly leads to the binding enviroment difference at the active sites of two isoforms,as represented by the electrostatic surface potential maps,which further results in the diverse dynamic behaviors of CBZ in two isoforms.Our research will help to the deep understanding of CYP isoform’s physilogical functions and open a door for the development of the isoform specific inhibitors.3.Molecular dynamics simulation of the interaction between azamulin with two CYP isoforms 3A4 and 3A5The unmarketed potential drug molecule azamurin has been found to be a specific inhibitor of CYP3A4 and CYP3A5 in recent years,but this molecule also shows different binding ability and activity to the two CYP3 A isomerases.In order to explore the microscopic mechanism,we used molecular dynamics simulation methods to study the dynamic interactions between CYP3A4 and CYP3A5 and azamulin.The simulation results showed that the binding of the same ligand resulted in different structural properties of the two proteins.First of all,compared with CYP3A4 without binding substrate,binding substrate azamulin can lead to flexibility changes in protein structure and increased flexibility in multiple peptide regions,that is,holo-CYP3A4 is more flexible than apo-CYP3A4.The structural changes of CYP3A5 are just the opposite.The binding of ligands enhances the rigidity of CYP3A5.Further,we analyzed in detail the representative structures of the stable phase during the kinetic simulation,and analyzed the detailed interactions between the ligand azamulin and two CYP3 A isomerases at the atomic level.It is speculated that the difference of composition and interaction of active sites is the fundamental cause of the change of structural properties of the two proteins. |
