Date Published: February 2, 2017
Publisher: Public Library of Science
Author(s): Kristen C. Witt, Luis Castillo-Menendez, Haitao Ding, Nicole Espy, Shijian Zhang, John C. Kappes, Joseph Sodroski, Jamie Kathleen Scott.
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.
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 [1–3]. The resulting mature Env complex consists of three gp120 exterior subunits and three gp41 transmembrane subunits. The metastable Env trimer is triggered to undergo conformational changes by binding to the receptors, CD4 and CCR5/CXCR4 [1,4–17]. CD4 binding induces the formation of a pre-hairpin intermediate, and CCR5/CXCR4 binding drives the formation of a stable gp41 six-helix bundle required for the fusion of the viral and target cell membranes [4–7,18–24].
Metastability is essential for the function of HIV-1 Env trimers. The high-potential-energy conformation of the unliganded Env is converted by receptor binding to lower-energy conformations on the virus entry pathway [1,5–7,10,11,18,21,85]. Even in its unliganded state, the Env from a laboratory-adapted HIV-1 strain has been shown to sample multiple conformations . Understanding the conformations of primary HIV-1 Env can facilitate approaches to treatment and prevention. For example, presenting the relevant conformations of Envs from transmitted/founder HIV-1 may be critical to the development of vaccines.