Structural basis of hypervariable region 1 dynamics on HCV envelope glycoproteins in immune evasion

The hypervariable region 1 (HVR1) of the hepatitis C virus E1E2 glycoprotein plays a crucial role in immune evasion through rapid mutation and shielding of conserved neutralizing epitopes. Despite its important role, the structural properties of HVR1 remain poorly understood, particularly how its conformation influences shielding mechanisms and receptor recognition. To date in all published E1E2 or E2 structures, the C-terminal part of HVR1 (394 to 411) remains unresolved. Using neutralizing antibody (nAb) RM1-38, which targets C-terminal HVR1 (residues 399 to 409), we investigated the folding, positioning, flexibility, and shielding function of the RM1-38 epitope, or antigenic site 399 (AS399), on HVR1 by X-ray crystallography and negative-stain electron microscopy (nsEM). The crystal structure of RM1-38 Fab with the HVR1 AS399 peptide, combined with mutagenesis analysis, delineated the molecular basis of its strain-specific recognition and neutralization. nsEM and competition enzyme-linked immunosorbent assay revealed that HVR1 exhibits pronounced conformational flexibility even in the presence of other nAbs bound to E1E2. Virus neutralization assays using a combination of RM1-38 and broadly neutralizing antibodies (bnAbs) demonstrated antagonistic effects of RM1-38 with the bnAbs. Furthermore, RM1-38 reduced E1E2 binding to cells expressing the scavenger receptor SR-BI, an HVR1-dependent receptor, but did not affect binding to CD81 tetraspanin. Together, these findings support a model in which HVR1 functions as a dynamic shield, where its conformational plasticity contributes to both receptor engagement and evasion of humoral immunity in persistent infection.