Ligand Recognition by the Lipid Transfer Domain of Human OSBP Is Important for Enterovirus Replication

Oxysterol-binding protein (OSBP), which trans-ports cholesterol and phosphatidylinositol 4-monophosphate (PtdIns[4]P) between different organelles, serves as a conserved host factor for the replication of various viruses, and OSBP inhibitors exhibit antiviral effects. Here, we determined the crystal structure of the lipid transfer domain of human OSBP in complex with endogenous cholesterol. The hydrocarbon tail and tetracyclic ring of cholesterol interact with the hydrophobic tunnel of OSBP, and the hydroxyl group of cholesterol forms a hydrogen bond network at the bottom of the tunnel. Systematic mutagenesis of the ligand-binding region revealed that M446W and L590W substitutions confer functional tolerance to an OSBP inhibitor, T-00127-HEV2. Employing the M446W variant as a functional replacement for the endogenous OSBP in the presence of T-00127-HEV2, we have identified previously unappreciated amino acid residues required for viral replication. The combined use of the inhibitor and the OSBP variant will be useful in elucidating the enigmatic in vivo functions of OSBP.

Automated summary

Oxysterol-binding protein (OSBP), as a conserved host factor, transports cholesterol and phosphatidylinositol 4-monophosphate (PtdIns[4]P) between different organelles, and serves as a factor for the replication of various viruses. In this study, the crystal structure of the lipid transfer domain of human OSBP in complex with endogenous cholesterol was determined, revealing the interaction between cholesterol and the hydrophobic tunnel of OSBP. Mutagenesis analysis identified specific amino acid residues required for viral replication, and the M446W variant showed tolerance to the OSBP inhibitor T-00127-HEV2, suggesting its potential use in studying the in vivo functions of OSBP.