Mapping the Binding\nHotspots and Transient Binding\nPockets on V‑Domain Immunoglobulin Suppressor of T Cell Activation\nProtein Surface

V-domain immunoglobulin suppressor of T cell activation\n(VISTA),\nan inhibitory immune checkpoint present on both immune and tumor cells,\nhas emerged as a highly promising target for cancer therapy due to\nits potential to overcome resistance encountered with existing immune\ncheckpoint treatments. VSIG-3 is determined as an inhibitory ligand\nfor VISTA, leading to the suppression of T cell proliferation. However,\nhotspots between VISTA/VSIG-3 protein–protein interaction remain\nambiguous, mainly attributed to the lack of the structure of the VISTA/VSIG-3\ncomplex. Therefore, in this study, in order to determine the energetic\ncontributions of the interfacial residues on VISTA, we first constructed\nVISTA/VSIG-3 complex models by the protein docking method, followed\nby molecular dynamics simulations, binding free-energy decomposition,\nand alanine scanning. Results suggested that the putative hotspots\nin VISTA comprise residues His32, Tyr37, Thr35, Glu47, Val48, Gln49,\nGlu53, Arg54, Gln73, His122, and His126. Moreover, the distribution\nof the hotspots was clustered into two regions (hot regions I and\nII), and by using the TRAPP tool, transient subpockets within the\nhot regions were identified. Furthermore, conformational states of\nthe binding pockets exhibiting druggability scores higher than those\nobserved in the crystal structure were found. Overall, we hope that\nthe findings outlined in this study can be used to facilitate the\ndevelopment of inhibitors targeting the VISTA/VSIG-3 immune checkpoint\npathway in the future.