The Neutralizing Face of Hepatitis C Virus E2 Envelope Glycoprotein

Abstract

The high genetic variability of hepatitis C virus, together with the high level of glycosylation on the viral envelope proteins shielding potential neutralizing epitopes, pose a difficult challenge for vaccine development. An effective hepatitis C virus (HCV) vaccine must target conserved epitopes and the HCV E2 glycoprotein is the main target for such neutralizing antibodies (NAbs). Recent structural investigations highlight the presence of a highly conserved and accessible surface on E2 that is devoid of N-linked glycans and known as the E2 neutralizing face. This face is defined as a hydrophobic surface comprising the front layer (FL) and the CD81 binding loop (CD81bl) that overlap with the CD81 receptor binding site on E2. The neutralizing face consists of highly conserved residues for recognition by cross-NAbs, yet it appears to be high conformationally flexible, thereby presenting a moving target for NAbs. Three main overlapping neutralizing sites have been identified in the neutralizing face: antigenic site 412 (AS412), antigenic site 434 (AS434), and antigenic region 3 (AR3). Here, we review the structural analyses of these neutralizing sites, either as recombinant E2 or epitope-derived linear peptides in complex with bNAbs, to understand the functional and preferred conformations for neutralization, and for viral escape. Collectively, these studies provide a foundation and molecular templates to facilitate structure-based approaches for HCV vaccine development.

Keywords: crystal structure; hepatitis C virus; neutralizing antibodies; neutralizing face; vaccine design.

Figure 1

The neutralizing face of the hepatitis C virus E2 glycoprotein. (A) Schematic representations of E2 regions (a.a. 384–746, based on the prototypic isolate H77 numbering system) colored by structural components with variable regions in gray, AS412 region in pink, front layer (FL) in cyan, β-sandwich in red, CD81 binding loop (CD81bl) in blue, back layer (BL) in green, and the stalk and transmembrane region (TM) in white. The a.a. sequence of the neutralization face (a.a. 412–446 and 525–535) and the epitope of mAb DAO5 is shown below. The AS412, antigenic region 3, and AS434 neutralization epitopes are marked in pink, dashed rectangle, and wheat. The epitope of the non-neutralizing mAb DAO5 is marked in green. The N-linked glycosylation sites surrounding the neutralizing face (N417, N423, N430, N532, and N540) are underlined. (B) Surface representation of the E2c structure (25) (PDB entry 4MWF) with the structural components colored as in (A). The neutralizing face is marked by a red dashed line. (C) The E2 neutralizing epitopes on the E2c structure. For the AS412 epitopes, a.a. 412–420 are modeled onto E2c based on the AP33 bNAb-AS412 crystal structure (PDB entry 4G6A). The conformational flexibility of AS412 related to the E2 (30) is schematically shown. The three known AS412 conformations (β-hairpin, semi-open, and open) for neutralization are shown on the right. (D) Summary of the E2 crystal structures. The two E2 core domain-mAb structures are marked by stars.