Journal Article

Substrate recognition and cleavage mechanism of the monkeypox virus core protease

Ji, Xiaoyun, Li, Dongxu, Wang, Wei, Cui, Wen, Cao, Junyuan, Zhang, Mingjing, Jia, Handi, Wu, Botao, Zhang, Xiaoyu, Yang, Xiuna, Yin, Yue, Xie, Xiong, Xiang, Yingchun, You, Tian, Yang, Kailin, Rao, Zihe, Wang, Haofeng, Jia, Menghan, Mao, Shengjiang, Duan, Yinkai, Nguyen, Henry C., Lan, Weiqi, Yang, Haitao, Dong, Xuxue, Shang, Weijuan, Sun, Zhanqi, Gao, Yan, Liu, Xiaoce, Liu, Hong, Hu, Shulei, Zhang, Yumin, Dai, Wenhao, Hu, Wanlong, Zhou, Hao, Zhang, Leike, Xu, Jin, Yang, Bei, Xiao, Gengfu

2025

Overview

Poxviruses cause severe diseases, including smallpox and mpox, that pose major threats to human health. The poxvirus core protease (Core Pro ) is essential for viral maturation and is highly conserved in poxviruses, making it an attractive antiviral target 1 . However, the structure of Core Pro remains unknown, hampering antiviral development. Here we determined the apo structure of monkeypox virus (MPXV) Core Pro and the structure of Core Pro in a complex with the inhibitor aloxistatin, a drug candidate for muscular dystrophy 2 . These structures show that Core Pro forms a homodimer that features a unique ‘dancing couple’ fold. The catalytic intermediate state of Core Pro was characterized by an aldehyde derivative from a natural substrate ( I-G18 ). This derivative binds covalently to the catalytic Cys328, shifting the active site of the viral protease from a closed conformation in the apo form to a favourable open conformation upon substrate binding. On the basis of the Core Pro – I-G18 complex, we designed a series of peptidomimetic inhibitors with a nitrile warhead, which could covalently anchor with the catalytic Cys328. These compounds inhibit Core Pro with half-maximal inhibitory concentrations of 44.9–100.3 nM, and exhibit potent and broad anti-poxvirus activity. Our studies provide a basis for designing wide-spectrum inhibitors against poxvirus infections. Structural and biochemical studies of the monkeypox virus core protease show that it exists as an active homodimer and indicate that the substrate-binding sites of core proteases are a promising target for antiviral drugs.

• Publisher: Nature Publishing Group UK,Nature Publishing Group
• Subject: 101/28 / 119/118 / 49/98 / 631/154 / 631/535 / 82/47 / 82/83 / Aldehydes / Aldehydes - chemistry / Aldehydes - metabolism / Antiviral Agents - chemistry / Antiviral Agents - pharmacology / Antiviral drugs / Biocatalysis - drug effects / Catalytic Domain - drug effects / Conformation / Crystallography, X-Ray / Design / Drug development / Efficiency / Humanities and Social Sciences / Humans / Inhibitors / Models, Molecular / Monkeypox virus - chemistry / Monkeypox virus - enzymology / Mpox / multidisciplinary / Oligopeptides - chemistry / Oligopeptides - metabolism / Oligopeptides - pharmacology / Peptidomimetics - chemistry / Peptidomimetics - metabolism / Peptidomimetics - pharmacology / Protease / Protease Inhibitors - chemistry / Protease Inhibitors - metabolism / Protease Inhibitors - pharmacology / Proteases / Proteinase / Proteins / Public health / Science / Science (multidisciplinary) / Severe acute respiratory syndrome coronavirus 2 / Smallpox / Substrate Specificity - drug effects / Viral diseases / Viral infections / Viral Proteases - chemistry / Viral Proteases - metabolism / Viral Proteins - antagonists & inhibitors / Viral Proteins - chemistry / Viral Proteins - metabolism / Viruses