Plasmodium proteases play both regulatory and effector roles in essential biological processes in this important pathogen and have long been investigated as drug targets. Plasmepsin V from P. falciparum (PfPMV) is an essential protease that processes proteins for export into the host erythrocyte and is a focus of ongoing drug development efforts. In the present study, recombinant protein production, inhibition assays, binding studies as well as molecular docking and molecular dynamics simulation studies were used to investigate the mode of binding of a PEXEL-based peptidomimetic and naphthoquinone compounds to PfPMV. Consistent with our previous study, refolded PfPMVs were produced with functional characteristics similar to the soluble counterpart. Naphthoquinone compounds inhibited PfPMV activity by 50% at 50 μM but did not affect pepsin activity. The IC₅₀ values of compounds 31 and 37 against PfPMV were 22.25 and 68.94 μM, respectively. Molecular dynamics simulations revealed that PEXEL peptide interacted with PfPMV active site residues via electrostatic interactions while naphthoquinone binding preferred van der Waal interactions. P₁′-Ser of the PfEMP2 substrate formed an additional H-bond with Asp365 promoting the catalytic efficiency. Additionally, the effect of metal ions on the secondary structure of PfPMV was examined. Our results confirmed that Hg²⁺ ions reversibly induced the changes in secondary structure of the protein whereas Fe³⁺ ions induced irreversibly. No change was observed in the presence of Ca²⁺ ions. Overall, the results here suggested that naphthoquinone derivatives may represent another source of antimalarial inhibitors targeting aspartic proteases but further chemical modifications are required.

疟原虫蛋白酶在该重要病原体的必需生物学过程中起调节作用和效应物作用，长期以来一直被作为药物靶标进行研究。恶性疟原虫（Pf PMV）的血浆蛋白酶V是一种必需的蛋白酶，其加工蛋白质以输出到宿主红细胞中，并且是正在进行的药物开发工作的重点。在本研究中，重组蛋白生产，抑制测定，结合研究以及分子对接和分子动力学模拟研究用于研究基于PEXEL的拟肽和萘醌化合物与Pf PMV的结合方式。与我们之前的研究一致，重新折叠Pf产生具有类似于可溶性对应物的功能特征的PMV。萘醌化合物在50μM时可抑制Pf PMV活性50％，但不影响胃蛋白酶活性。化合物31和37对Pf PMV的IC ₅₀值分别为22.25和68.94μM。分子动力学模拟显示，PEXEL肽通过静电相互作用与Pf PMV活性位点残基相互作用，而萘醌结合则更倾向于范德华相互作用。P ₁ '的的-Ser Pf的EMP2衬底上形成一个附加的H-键与Asp365促进催化效率。此外，金属离子对Pf二级结构的影响检查了PMV。我们的结果证实，Hg ²⁺离子可逆地诱导蛋白质二级结构的变化，而Fe ³⁺离子则不可逆地诱导。在Ca ²⁺离子的存在下未观察到变化。总的来说，这里的结果表明萘醌衍生物可能代表了靶向天冬氨酸蛋白酶的抗疟疾抑制剂的另一种来源，但是还需要进一步的化学修饰。