Both lipid environment and pH are critical for determining physiological solution structure of 3‐D‐conserved epitopes of the HIV‐1 gp41‐MPER peptide P1

Abstract

In terms of background, the solution structure of monomeric peptide P1 (residues 649-683), located in the conserved membrane proximal region (MPER) of HIV-1 envelope glycoprotein gp41, is first reported here in dodecylphosphocholine (DPC) micelles. P1 is the minimal MPER region that permits interaction with the mucosal galactosyl ceramide HIV-receptor; it also contains epitopes recognized by major gp41-specific, broadly neutralizing immunoglobulin Gs (IgGs), 2F5 and 4E10, determinant in HIV fusion/infection. Our principal findings were as follows: the structural stability of P1 is pH dependent, as the alpha-helix comprising Q653 I682, present at pH 3.3, is destabilized at higher pH values. At pH 6, the E-rich N-terminal half of P1 (residues 650-666), partially overlapping the 2F5-specific epitope, becomes fully disordered, while the W-rich C-terminal half conserves two shorter helices (W666-W670 and W672-W680), separated by a well-defined bend overlapped by the 4E10-specific epitope. The two IgGs bind to P1 on DPC micelles with binding parameters (K(d)) in the nanomolar range. Next, P1 was derivatized with phosphatidylethanolamine at its C terminal and inserted into liposomes of varied lipid composition, thereby enabling P1 to move laterally. Alternatively, an infectious virus-binding assay was established. The K(d) of both 2F5 and 4E10 IgGs measured on viral liposome and virus are similar and much lower than for the binding of the free peptide. In conclusion, P1, in a lipid environment, is an optimized MPER-derived peptide suitable for designing an immunogen inducing broadly neutralizing antibodies to HIV.

Stable Docking of Neutralizing Human Immunodeficiency Virus Type 1 gp41 Membrane-Proximal External Region Monoclonal Antibodies 2F5 and 4E10 Is Dependent on the Membrane Immersion Depth of Their Epitope Regions

“…However, more defined secondary structures have been described for some shorter peptides (D 664 -K 683 ) both in solution and in micelle-or liposomebound states in infrared (17,26,30) and CD spectroscopic studies (16). More recently, nuclear magnetic resonance spectroscopic studies of micelle-bound peptides corresponding to E 662 -K 683 (31) and S 649 -K 683 (8) revealed that the C-terminal half of MPER consists of two shorter helices separated by a short hinge and that the N-terminal half of MPER is mostly disordered.…”

“…MPER peptide anchorage on the membrane surface facilitates efficient 2F5 and 4E10 binding. Recent studies that have described the binding of MAbs 2F5 and 4E10 to MPER peptides on liposomes used either no membrane anchor (31) or a lipidic anchor (8) and relied on the inherent membrane affinity of the MPER peptides. The importance of membrane anchoring on peptide orientation and the effect on MAb binding have not been studied.…”

“…Thus, a key question that needed to be addressed was whether the MPER peptides adopted a unique structure in the membrane environment that favored the docking of 2F5 and 4E10. Even though highresolution structures of MPER peptides of various lengths in micelles or bicelles have been reported (8,31), no high-resolution structure of membrane-bound MPER peptide is available except for a structure based on the electron paramagnetic resonance-measured membrane immersion depth parameters of MPER amino acid residues (31). The CD spectroscopic experiments showed that the nominal epitope and biepitope peptides adopted ordered structures with different helical contents (Table 4).…”

Structure of an HIV-1–neutralizing antibody target, the lipid-bound gp41 envelope membrane proximal region trimer

“…The short peptides were either solubilized in detergent micelles (39,44,45) or in bound forms cocrystallized with MPER-recognizing antibodies (31,32). Of the trimeric structures, one was not membrane-associated and does not bind 2F5 or 4E10 antibodies (38), and the other was part of a six-helix bundle representing the postfusion state of gp41 (13).…”

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

“…2-4A). Structures of shorter or even longer peptides show spots of partial structuring as 3 10 -and ␣-helix (33,35,73,(75)(76)(77), but none of them display continuous helical structures for the sequence spanning the full 2F5 epitope plus the downstream aromatic rich sequence preceding the TMD anchor (Fig. 1).…”