Catalytic mechanism and inhibition of Plasmodium sp. serine hydroxymethyltransferase by computational simulations

Abstract

Malaria has spread in many countries with a 12% increase in death cases after the COVID-19 pandemic. Over the Greater Mekong subregion, the clinical efficacy of antimalarial drugs has been reduced due to resistance. Target-based approaches on attractive drug targets, such as Plasmodium serine hydroxymethyltransferases (SHMTs) exhibiting distinct structure and function as well as kinetic mechanisms from the human enzyme homologue, are highly useful methods to be used for bypassing the present resistance in the field. In this study, the catalytic mechanism of Plasmodium SHMT was investigated following the retro-aldol proposed scheme by the combined quantum mechanics/molecular mechanics molecular dynamics (QM/MM MD) simulations. E56 and Y64, residues in the Plasmodium SHMT binding pocket, were suggested as a general base and general acid. The PM6 and AM1/D levels of theory were applied to the QM region. However, the reaction was stopped after the formaldehyde occurred. This result indicated the unfavorable of the level of theory on the QM region. Further study should be performed by another method e.g., QUICK package in AMBER20 for HF and DFT calculations. Additionally, the 500-ns MD simulations were carried out to investigate the mode of action of pyrazolopyran(+)-85 and pyrazolopyran(+)-86 with the most attractive inhibition efficiency in Plasmodium falciparum and P. vivax SHMTs. The results suggested that the binding affinity of pyrazolopyran(+)-86 is more favorable than pyrazolopyran(+)-86 (~ 2 kcal/mol). L124, G128, H129, L130, K139, N356, and T357 are essential residues for inhibitor binding. The rational structure-based drug design suggested that the isopropyl moiety on the pyrazolopyran core should be changed to the negatively charged group (e.g., carboxylate group) for interacting with the positively charged residue R371. Alternatively, the phenolic compounds could be substituted with a phenyl or piperidine ring to promote hydrogen bond formation with the surrounding residues. Moreover, to find other inhibitors with a rapid method, drug repurposing was performed. The FDA-approved drugs were screened over seven Plasmodium SHMT structures. The result revealed that only eight potent compounds exhibited the lowest binding energy in all complexes and were then investigated by the enzyme-based assay. The experimental results indicated the lowest IC50 value (106 ± 1µM) was observed on PfSHMT by amphotericin B. The binding affinity of amphotericin B (–11.15 ± 0.09 kcal/mol) was in the same range as pyrazolopyran-based inhibitors. Moreover, Y63, L124, L130, F134, V141, K251, D258, N259, and S263 were the key binding residues to amphotericin B. These findings could provide insights into the mode of inhibition of pyrazolopyran-based inhibitors and introduce a new core structure of inhibitor that differs from the previous SHMT inhibitors which could be a rational idea for novel antimalarial drug design.