Abstract
The entry of human immunodeficiency virus (HIV-1) into host cells is mediated by the viral envelope glycoproteins (Envs), which are derived by the proteolytic cleavage of a trimeric gp160 Env precursor. The mature Env trimer is a major target for entry inhibitors and vaccine-induced neutralizing antibodies. Env interstrain variability, conformational flexibility and heavy glycosylation contribute to evasion of the host immune response, and create challenges for structural characterization and vaccine development. Here we investigate variables associated with reconstitution of the HIV-1 Env precursor into nanodiscs, nanoscale lipid bilayer discs enclosed by membrane scaffolding proteins. We identified detergents, as well as lipids similar in composition to the viral lipidome, that allowed efficient formation of Env-nanodiscs (Env-NDs). Env-NDs were created with the full-length Env precursor and with an Env precursor with the majority of the cytoplasmic tail intact. The self-association of Env-NDs was decreased by glutaraldehyde crosslinking. The Env-NDs exhibited an antigenic profile expected for the HIV-1 Env precursor. Env-NDs were recognized by broadly neutralizing antibodies. Of note, neutralizing antibody epitopes in the gp41 membrane-proximal external region and in the gp120:gp41 interface were well exposed on Env-NDs compared with Env expressed on cell surfaces. Most Env epitopes recognized by non-neutralizing antibodies were masked on the Env-NDs. This antigenic profile was stable for several days, exhibiting a considerably longer half-life than that of Env solubilized in detergents. Negative selection with weak neutralizing antibodies could be used to improve the antigenic profile of the Env-NDs. Finally, we show that lipid adjuvants can be incorporated into Env-NDs. These results indicate that Env-NDs represent a potentially useful platform for investigating the structural, functional and antigenic properties of the HIV-1 Env trimer in a membrane context.
Highlights
Human immunodeficiency virus (HIV-1) entry into the host cell is mediated by the viral envelope glycoproteins (Envs), which are derived by the proteolytic cleavage of a trimeric gp160 Env precursor [1,2,3]
To increase the stability of Env, these studies were carried out using the uncleaved Env precursors, Env(-) and Env(-)Δ808 glycoproteins, produced in Chinese hamster ovary (CHO) cells [79]
When purified HIV-1JR-FL Env(-)Δ808 trimers and brain lipids in CHAPSO were mixed with the membrane scaffolding protein MSP1E2 and the detergent concentration was subsequently reduced by incubation with hydrophobic beads, complexes at a size expected for Env-NDs were observed by size-exclusion chromatography (Fig 1B)
Summary
Human immunodeficiency virus (HIV-1) entry into the host cell is mediated by the viral envelope glycoproteins (Envs), which are derived by the proteolytic cleavage of a trimeric gp160 Env precursor [1,2,3]. The Env trimer is the sole virus-specific target on the virion surface available for the binding of neutralizing antibodies [1,2,3,25,26,27,28]. To allow persistence in the host, HIV-1 Envs have evolved features to minimize the elicitation and impact of broad neutralizing antibodies [1,29,30]. These features include surface variability among HIV-1 strains, conformational lability, and a heavy glycan coat [29,30,31,32,33,34]. After several years of infection and in a minority of infected individuals are broadly neutralizing antibodies generated [44,45,46,47,48,49]
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