Discovery of a Heparan Sulfate Binding Domain in Monkeypox Virus H3 as an Anti-poxviral Drug Target Combining AI and MD Simulations

Viral adhesion to host cells is a critical step in infection for many viruses, including monkeypox virus (MPXV). In MPXV, the H3 protein mediates viral adhesion through its interaction with heparan sulfate (HS), yet the structural details of this interaction have remained elusive. Using AI-based structural prediction tools and molecular dynamics (MD) simulations, we identified a novel, positively charged α-helical domain in H3 that is essential for HS binding. This conserved domain, found across orthopoxviruses , was experimentally validated and shown to be critical for viral adhesion, making it an ideal target for antiviral drug development. Targeting this domain, we designed a protein inhibitor, which disrupted the H3-HS interaction, inhibited viral infection in vitro and viral replication in vivo , offering a promising antiviral candidate. Our findings reveal a novel therapeutic target of MPXV, demonstrating the potential of combination of AI-driven methods and MD simulations to accelerate antiviral drug discovery.