Trimerization of the HIV Transmembrane Domain in Lipid Bilayers Modulates Broadly Neutralizing Antibody Binding

Abstract

The membrane-proximal external region (MPER) of HIV gp41 is an established target of antibodies that neutralize a broad range of HIV isolates. To evaluate the role of the transmembrane (TM) domain, synthetic MPER-derived peptides were incorporated into lipid nanoparticles using natural and designed TM domains, and antibody affinity was measured using immobilized and solution-based techniques. Peptides incorporating the native HIV TM domain exhibit significantly stronger interactions with neutralizing antibodies than peptides with a monomeric TM domain. Furthermore, a peptide with a trimeric, three-helix bundle TM domain recapitulates the binding profile of the native sequence. These studies suggest that neutralizing antibodies can bind the MPER when the TM domain is a three-helix bundle and this presentation could influence the binding of neutralizing antibodies to the virus. Lipid-bilayer presentation of viral antigens in Nanodiscs is a new platform for evaluating neutralizing antibodies. KeywordsPeptides; Membrane Proteins; Nanostructures; Antibodies; HIV The membrane proximal external region (MPER) is a portion of the envelope glycoprotein gp41 on the surface of HIV virions. The MPER is critical to membrane fusion and infection, and is one of the most highly conserved regions of HIV [2] . A number of broadly neutralizing antibodies including 4E10, Z13e1, 2F5, and 10E8 have been identified that target overlapping regions of the MPER, and a broad analysis of patient sera suggest that MPER-directed antibodies are more common than previously thought [3] . Neutralizing antibodies identified from HIV+ individuals have been shown to provide protection against Correspondence to: Philip E. Dawson. HHS Public Access Author ManuscriptAuthor Manuscript Author ManuscriptAuthor Manuscript intravenous and mucosal challenge in passive transfer animal studies, suggesting that a neutralizing antibody response could protect against HIV [4] . Despite the apparent linear nature of described MPER epitopes, the design of effective MPER immunogens has had limited success [5] . Thus the MPER is ripe for a reverse vaccinology approach to elucidate the interactions between known broadly neutralizing antibodies and their epitopes in order to direct the iterative design of immunogens.The structure of the MPER is largely unknown in the context of Env trimers, despite highresolution structural information regarding Env trimers both on the virion surface and in soluble forms. Cryo-EM data of virion surfaces show low density and low signal to noise at the MPER, suggesting less protein mass or greater disorder in this region [6] . Recent cryo-EM [1] and x-ray crystallography structures [7] of stabilized soluble gp41-gp120 trimers required removal of the MPER to obtain stable constructs. The conformation of the MPER is still an open question of great interest and import to antigen design and general understanding of HIV.Recognizing the importance of the membrane in understanding the MPER, the MPER has been presented in association ...

Nanodiscs in Membrane Biochemistry and Biophysics

“…223 Based on the fact that the presence of membrane and formation of trimer are necessary for efficient binding with neutralizing antibodies, a structural model of MPER fragment was suggested. 223 …”

“…A recent publication described the success using the HIV envelope spike proteins and their fragments. 90,223 The small size of Nanodiscs and their stability in plasma improved antigen life-time in vivo and may prove to be a most useful approach in vaccine development. Viability of this concept is supported by the recent successful production of antibodies in phage display for several proteins that were self-assembled in Nanodiscs.…”

The power, pitfalls and potential of the nanodisc system for NMR-based studies

“…Sodium cholate has been shown to be an efficient detergent for phospholipid solubilization (Bayburt and Sligar, 2010) but sodium dodecylsulfate (SDS) or other detergents can be used as well (D'Antona et al, 2014). In some cases the same detergent can be used for resuspending the lipids and solubilizing the membrane proteins Reichart et al, 2016), e.g. when SDS-solubilized bacterioopsin is directly refolded into NDs , but often milder detergents are used to directly transfer a folded protein conformation into the NDs.…”

HIV-1 Env gp41 Transmembrane Domain Dynamics Are Modulated by Lipid, Water, and Ion Interactions

“…1) is anchored in a cholesterol-rich lipid bilayer (21) that is flanked by the membrane proximal external region (MPER) on the exofacial leaflet and the cytoplasmic tail (CT) on the cytofacial leaflet. In the prefusion state, either one (22) or three (20,23,24) single-pass a-helices span the membrane (Fig. 1 A).…”