HIV gp41-Antibody Interaction at the Viral Membrane Interface Defined by EPR Spectroscopy

Abstract

HIV enters human T cells through the fusion of viral and host-cell membranes. This fusion process is mediated by a surface protein, gp41, and the platform provided by the cholesterol-rich viral membrane. The membrane proximal ectodomain region (MPER) of gp41 plays a critical role in this fusion process and is a major target of anti-gp41 antibodies and vaccine design. Here, EPR and NMR techniques were used to define MPER structure on the membrane, and how neutralizing anti-gp41 antibodies recognize their membrane-immersed epitopes and disrupt a hinge-related function of the MPER. The analyses of several HIV-1 clade B and clade C MPERs revealed a structurally conserved pair of helices immersed in the viral membrane separated by a flexible hinge, which include critical helix capping residues. Double alanine mutations of the capping residues result in an altered hinge structure with a deeper lipid-buried MPER middle region, as well as reduced viral fusion and infectivity. Furthermore, neutralizing anti-gp41 antibodies disrupt the MPER hinge function by perturbing MPER hinge orientation, and/or extracting part of the MPER from the membrane. The interaction can be a stepwise rearrangement through an apparent scoop-like movement of the antibodies’ long and unique CDRH3 segments. Mutations of the CDRH3 segments reduced the ability of the antibodies to extract MPER residues from the membrane, without affecting peptide binding in solution. In addition, MPER-membrane interaction and antibody binding are modulated by lipid composition and cholesterol content. These findings have revealed important features of gp41-antibody interaction at the viral membrane interface.