2. Academic Validation
  3. Hepatitis E Virus ORF1 Polyprotein Harbors a Pocket-Like Cavity That Is Vital for Virus Replication and Represents a Novel Antiviral Target

Hepatitis E Virus ORF1 Polyprotein Harbors a Pocket-Like Cavity That Is Vital for Virus Replication and Represents a Novel Antiviral Target

  • Adv Sci (Weinh). 2025 Oct 2:e01699. doi: 10.1002/advs.202501699.
Xiaohui Ding 1 2,  Dou Zeng 1 2,  Dan Liu 1 2,  Yingying Bian 1 2,  Bin Li 1 2,  Zheng Li 1 2,  Qiudi Li 1 2,  Shiquan Liang 1 2,  Yunlong Si 3,  Qili Yao 1 2,  Yibo Ding 1 2,  Jiahui Zhu 3,  Xiangyang Li 1 2,  Kuiyang Zheng 1 2,  Hongbo Guo 1 2,  Wenshi Wang 1 2
Affiliations

Affiliations

  • 1 Department of Pathogen Biology and Immunology, School of Basic Medical Sciences, Xuzhou Medical University, Xuzhou, 221004, China.
  • 2 Jiangsu Key Laboratory of Immunity and Metabolism, Jiangsu International Laboratory of Immunity and Metabolism, Xuzhou Medical University, Xuzhou, 221004, China.
  • 3 Jiangsu Key Laboratory of Brain Disease and Bioinformation, Research Center for Biochemistry and Molecular Biology, Xuzhou Medical University, Xuzhou, 221004, China.
PMID: 41038801 DOI: 10.1002/advs.202501699
Abstract

Hepatitis E virus (HEV) is the leading cause of acute viral hepatitis worldwide, yet no FDA-approved anti-HEV medication available. Elucidating HEV replication machinery is therefore crucial for identifying novel Antiviral targets, and consequently developing potent antivirals. The nonstructural ORF1 polyprotein is pivotal for HEV replication. Herein, it is revealed that the ORF1 X domain is a critical component of HEV replication machinery. Interestingly, the ADP-ribose hydrolase activity of X domain per se is dispensable for HEV replication. Instead, the X domain supports HEV replication through its interdomain interaction with the RNA-dependent RNA polymerase (RdRp). Structure-based functional analysis reveals that X and RdRp jointly create a "pocket-like" cavity (PC) at their interaction interface. Site-directed mutagenesis disrupting the PC integrity completely abolishes HEV replication, demonstrating its crucial role in the viral life cycle. Through a PC-targeted high-throughput in silico screening pipeline, combined with molecular docking analysis, surface plasmon resonance assays, and advanced in vitro HEV models, saikosaponin D and liriopesides B are identified as potent HEV inhibitors targeting this critical interface. Collectively, this study identifies a novel structure within ORF1 polyprotein that is crucial for HEV replication, and demonstrates the feasibility of developing novel antivirals by targeting this PC structure.

Keywords

ORF1 polyprotein; RNA‐dependent RNA polymerase; X domain; antiviral target; hepatitis E virus; replication machinery.

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