Crystallographic Definition of the Epitope Promiscuity of the Broadly Neutralizing Anti-Human Immunodeficiency Virus Type 1 Antibody 2F5: Vaccine Design Implications

Abstract

The quest to create a human immunodeficiency virus type 1 (HIV-1) vaccine capable of eliciting broadly neutralizing antibodies against Env has been challenging. Among other problems, one difficulty in creating a potent immunogen resides in the substantial overall sequence variability of the HIV envelope protein. The membrane-proximal region (MPER) of gp41 is a particularly conserved tryptophan-rich region spanning residues 659 to 683, which is recognized by three broadly neutralizing monoclonal antibodies (bnMAbs), 2F5, Z13, and 4E10. In this study, we first describe the variability of residues in the gp41 MPER and report on the invariant nature of 15 out of 25 amino acids comprising this region. Subsequently, we evaluate the ability of the bnMAb 2F5 to recognize 31 varying sequences of the gp41 MPER at a molecular level. In 19 cases, resulting crystal structures show the various MPER peptides bound to the 2F5 Fab. A variety of amino acid substitutions outside the 664 DKW 666 core epitope are tolerated. However, changes at the 664 DKW 666 motif itself are restricted to those residues that preserve the aspartate's negative charge, the hydrophobic alkyl-stacking arrangement between the ␤-turn lysine and tryptophan, and the positive charge of the former. We also characterize a possible molecular mechanism of 2F5 escape by sequence variability at position 667, which is often observed in HIV-1 clade C isolates. Based on our results, we propose a somewhat more flexible molecular model of epitope recognition by bnMAb 2F5, which could guide future attempts at designing small-molecule MPER-like vaccines capable of eliciting 2F5-like antibodies.

Neutralizing Epitopes in the Membrane-Proximal External Region of HIV-1 gp41 Are Influenced by the Transmembrane Domain and the Plasma Membrane

“…4A), supporting previous results in an ELISA-based system of detection (55), and viral neutralization studies (65), and in agreement with structural analysis (9). Replacement of Trp672 and Phe673 with Ala decreased binding by 4E10 (P Ͻ 0.05) (Fig.…”

Ablation of the Complementarity-Determining Region H3 Apex of the Anti-HIV-1 Broadly Neutralizing Antibody 2F5 Abrogates Neutralizing Capacity without Affecting Core Epitope Binding

“…The competitive ELISA measurements using core peptides as competitors also showed no difference in relative binding affinities for the different 2F5 Fab constructs. These results are in agreement with the structural data that show that the molecular interaction between the 2F5 paratope and core gp41 epitope residues is mediated by 2F5 amino acids other than those at the apex of the CDR H3 loop (6,24,37). Altogether, we believe that the present neutralization and binding affinity results represent strong evidence indicating that the 2F5 CDR H3 apex residues are essential for neutralization but are not involved in core epitope binding.…”

“…One of these antibodies, bnAb 2F5, has been extensively characterized in an effort to determine the molecular mechanism underlying its broad neutralization capability and with the objective of guiding vaccine immunogen design efforts. Although the core recognition site of 2F5 has been mapped to the 662 ELDKWAS 669 MPER linear sequence of gp41 and the atomic interactions of the core antigenic complex have been described in detail (6,24,37), the exact mechanism of broad HIV-1 neutralization by 2F5 remains elusive. Fab constructs for a glycosylated gp41 construct spanning residues 541 to 682 and a nonglycosylated gp41 construct spanning residues 535 to 669 were performed.…”

“…The interaction between 2F5 and its minimal linear epitope ( 662 ELDKWAS 668 ) has been extensively characterized in numerous structural studies (6,24,37). Briefly, the primary gp41 epitope as recognized by 2F5 assumes a ␤-turn conformation, with core residues 664 DKW 666 in the center of the turn.…”

Structure and Immunogenicity of a Peptide Vaccine, Including the Complete HIV-1 gp41 2F5 Epitope

“…The presence of these structures would be consistent with the flexibility found in the case of shorter peptides (35). Moreover, they would be consistent with the type I ␤-turn adopted by the 664 DKW 666 core epitope sequence in Fab2F5-peptide complexes (14,36,78). In contrast, our NMR structures did not display the extended conformation found in the crystal structures of those complexes for the preceding N-terminal residues 656 NEQELLEL 663 (14,36).…”