Periodic Reporting for period 1 - metabolicomp (Computational dynamics studies of drug metabolism by P450 enzymes)
Reporting period: 2017-06-21 to 2019-06-20
Basic mechanisms of desaturation and hydroxylation of ethylcarbamate by P450 model active site have been studied computationally; however, non-statistical dynamics had not been explored. After elucidating the stationary points along the PES, we carried out dynamics study with ethylcarbamate substrate to examine how the flat potential energy surfaces following C-H abstraction may play a role in product distributions. The aims of the project are to develop an understanding for alcohol/alkene product selectivity for ethylcarbamate using quasi-classical dynamics simulations, and to also understand the difference between heme and non-heme Fe-containing active site in product selectivity. In an extended effort to study reactive intermediates and natural metabolites, we collaborated with Burton group in Oxford to study oxonium ions and their roles in natural product biosyntheses.
To understand the difference between heme and non-heme enzymes, we needed to further examine the reactions catalyzed by non-heme active site. Potential energy surfaces were elucidated for the stepwise N-demethylation reactions for KMe3 and KMe3 methylated lysine with (AcO)2(imidazole)2Fe=O. Transition state structures and corresponding minima were computed for quintet and triplet surfaces, using the same method as above. Quasi-classical dynamics were then simulated for both substrates, starting from the hydrogen abstraction transition state. 100 trajectories were simulated for each system. Comparing the KMe3 and KMe2 system, an additional side product was observed for the KMe2 substrate due to the presence of the NH proton. This prompted reexamination of the potential energy surface with DFT for the formation of this side product. The results are discussed in more details in the upcoming publication.
Following the idea of site selectivity in enzyme active site and how that would affect product distribution, we collaborated with an experimental group in the same department to investigate the role of oxonium ions as reactive intermediates in several natural product biosynthesis. I performed quantum mechanical calculations to help confirm the identity of the oxonium ion intermediates by sampling various conformations of oxonium ions, computing chemical shifts for the ensemble of energetically relevant conformers, and then comparing the computational NMR data to their experimental data. This workflow was repeated for all 5 oxonium ions. Transition state structures (TSSs) were calculated for 1,2-hydride shifts to help understand the decomposition of the reactive oxonium ion. TSSs were also calculated for the reactions of forming oxonium ions and for the reactions between oxonium ions and nucleophiles. Though there are three possible sites on the oxonium ions for nucleophilic attack, the observed product distribution in earlier experimental study suggested that one site was more favorable. Distortion-interaction analyses revealed that reaction at this site involved the least structural reorganization of the substrate in enzyme active site. Manuscript with detailed discussion of these results are under reviews for publication.