Insights into red sea bream iridovirus pathogenesis: Unveiling host-pathogen interactions using membrane yeast...

Red sea bream iridovirus (RSIV), a member of the genus Megalocytivirus, poses a substantial threat to global aquaculture through its infection of marine fish species. The major capsid protein (MCP) represents the principal immunogenic component of RSIV, yet the mechanisms governing its pathogenicity is unclear. Here, we employed a comprehensive approach combining protein interaction screening with artificial intelligence-based structural prediction to elucidate the interface between RSIV and its host. In brief, by constructing a yeast cDNA library of membrane protein in red snapper and using a membrane yeast two hybrid system, a cDNA library targeting RSIV-MCP was screened, and 53 different host interaction partners were identified from the preliminary screening of 96 positive clones. Reverse validation confirmed robust protein-protein interactions for all candidates, including heat shock proteins Hsc70, HSP90β, and HSC71-like protein. Leveraging AlphaFold3 for structural prediction, we generated high-confidence models of MCP and HSC70, enabling molecular docking and dynamics simulations. The resulting complex demonstrated structural stability within 100 ns, as reflected by convergence of root-mean-square deviation and radius of gyration values. Computational analyses revealed an extensive hydrogen-bond network at the binding interface. This finding is further supported by a calculated binding free energy of -318.45 kJ/mol, indicating a potentially strong interaction between MCP and HSC70. Mutational analysis identified key residues 646ILE and 452ILE as critical mediators of complex formation, with hydrogen bonding and van der Waals interactions serving as primary stabilizing forces. These results provide insights into RSIV-host protein interactions, establishing a foundation for understanding viral pathogenesis and developing targeted therapeutic strategies against this economically significant aquatic pathogen.