The molecular mechanism of the proteolysis reaction catalyzed by SARS-CoV-2 M^pro, one of the enzymes essential for the replication process of the virus responsible for the COVID-19 pandemic, is described using computational QM/MM methods.

In this tutorial review we show how the methods and techniques of computational chemistry have been applied to the understanding of the physical basis of the rate enhancement of chemical reactions by enzymes. This is to answer the question: Why is the activation free energy in enzyme catalysed reactions smaller than the activation free energy observed in solution? Two important points of view are presented: Transition State (TS) theories and Michaelis Complex (MC) theories. After reviewing some of the most popular computational methods employed, we analyse two particular enzymatic reactions: the conversion of chorismate to prephenate catalysed by Bacillus subtilis chorismate mutase, and a methyl transfer from S-adenosylmethionine to catecholate catalysed by catechol O-methyltransferase. The results and conclusions obtained by different authors on these two systems, supporting either TS stabilisation or substrate preorganization, are presented and compared. Finally we try to give a unified view, where a preorganized enzyme active site, prepared to stabilise the TS, also favours those reactive conformations geometrically closer to the TS.

The environmental problems derived from the generalized plastic consumption and disposal could find a friendly solution in enzymatic biodegradation. Recently, two hydrolases from Ideonella sakaiensis 201-F6 and the metagenome-derived leaf-branch compost cutinase (LCC), more specially the improved ICCG variant, have revealed degradation activity toward poly ethylene terephthalate (PET). In the present study, the reaction mechanism of this polymer breakage is studied at an atomic level by multiscale QM/MM molecular dynamics simulations, using semiempirical and DFT Hamiltonians to describe the QM region. The obtained free energy surfaces confirmed a characteristic four-step path for both systems, with activation energies in agreement with the experimental observations. Structural analysis of the evolution of the active site along the reaction progress and the study of electrostatic effects generated by the proteins reveal the similarity in the behavior of the active site of these two enzymes. The origin of the apparent better performance of the LCC-ICCG protein over PETase must be due to its capabilities of working at higher temperature and its intrinsic relationship with the crystallinity grade of the polymer. Our results may be useful for the development of more efficient enzymes in the biodegradation of PET.

The SARS-CoV-2 main protease (M^pro) is essential for replication of the virus responsible for the COVID-19 pandemic, and one of the main targets for drug design. Here, we simulate the inhibition process of SARS-CoV-2 M^pro with a known Michael acceptor (peptidyl) inhibitor, N3. The free energy landscape for the mechanism of the formation of the covalent enzyme-inhibitor product is computed with QM/MM molecular dynamics methods. The simulations show a two-step mechanism, and give structures and calculated barriers in good agreement with experiment. Using these results and information from our previous investigation on the proteolysis reaction of SARS-CoV-2 M^pro, we design two new, synthetically accessible N3-analogues as potential inhibitors, in which the recognition and warhead motifs are modified. QM/MM modelling of the mechanism of inhibition of M^pro by these novel compounds indicates that both may be promising candidates as drug leads against COVID-19, one as an irreversible inhibitor and one as a potential reversible inhibitor.

Significance: The role of protein dynamics in enzyme catalysis remains a topic of considerable debate. Here, we use a combination of experimental and computational methods to identify the origins of the observed changes in reactivity on isotopic substitution of dihydrofolate reductase fromEscherichia coli. Isotopic substitution causes differences in environmental coupling to the hydride transfer step and protein dynamics have therefore a small but measurable effect on the chemical reaction rate.