School of Life Sciences, Tianjin University, Tianjin, China.
Tianjin International Joint Academy of Biotechnology and Medicine, Tianjin, China; State Key Laboratory of Medicinal Chemical Biology and College of Pharmacy, Nankai University, Tianjin, China.
Biochem Biophys Res Commun. 2019 Apr 16;511(4):794-799. doi: 10.1016/j.bbrc.2019.02.105. Epub 2019 Mar 2.
Mouse hepatitis virus A59 (MHV-A59) is a representative member of the genus betacoronavirus within the subfamily Coronavirinae, which infects the liver, brain and respiratory tract. Through different inoculation routes, MHV-A59 can provide animal models for encephalitis, hepatitis and pneumonia to explore viral life machinery and virus-host interactions. In viral replication, non-structural protein 5 (Nsp5), also termed main protease (M), plays a dominant role in processing coronavirus-encoded polyproteins and is thus recognized as an ideal target of anti-coronavirus agents. However, no structure of the MHV-A59 M has been reported, and molecular exploration of the catalysis mechanism remains hindered. Here, we solved the crystal structure of the MHV-A59 M complexed with a Michael acceptor-based inhibitor, N3. Structural analysis revealed that the Cβ of the vinyl group of N3 covalently bound to C145 of the catalytic dyad of M, which irreversibly inactivated cysteine protease activity. The lactam ring of the P1 side chain and the isobutyl group of the P2 side chain, which mimic the conserved residues at the same positions of the substrate, fit well into the S1 and S2 pockets. Through a comparative study with M of other coronaviruses, we observed that the substrate-recognition pocket and enzyme inhibitory mechanism is highly conservative. Altogether, our study provided structural features of MHV-A59 M and indicated that a Michael acceptor inhibitor is an ideal scaffold for antiviral drugs.
鼠肝炎病毒 A59（MHV-A59）是冠状病毒科冠状病毒亚科β冠状病毒属的代表性成员，可感染肝脏、大脑和呼吸道。通过不同的接种途径，MHV-A59 可为脑炎、肝炎和肺炎提供动物模型，以探索病毒的生命机制和病毒-宿主相互作用。在病毒复制过程中，非结构蛋白 5（Nsp5），也称为主要蛋白酶（M），在加工冠状病毒编码的多蛋白中起主导作用，因此被认为是抗冠状病毒药物的理想靶标。然而，尚未报道 MHV-A59 M 的结构，对其催化机制的分子探索仍然受到阻碍。在这里，我们解决了与迈克尔受体基抑制剂 N3 结合的 MHV-A59 M 的晶体结构。结构分析表明，N3 的乙烯基的 Cβ与 M 的催化二联体的 C145 共价结合，不可逆地使半胱氨酸蛋白酶失活。P1 侧链的内酰胺环和 P2 侧链的异丁基，模拟底物相同位置的保守残基，很好地适合 S1 和 S2 袋。通过与其他冠状病毒的 M 进行比较研究，我们观察到底物识别口袋和酶抑制机制高度保守。总之，我们的研究提供了 MHV-A59 M 的结构特征，并表明迈克尔受体抑制剂是抗病毒药物的理想支架。
