Topological Analysis of HIV-1 Glycoproteins Expressed In Situ on Virus Surfaces Reveals Tighter Packing but Greater Conformational Flexibility than for Soluble gp120

Abstract

bIn natural infection, antibodies interact with HIV-1 primarily through nonfunctional forms of envelope glycoproteins (Env), including uncleaved (UNC) gp160 and gp41 stumps. These antigens are important to fully characterize, as they may be decoys that promote nonneutralizing responses and may also be targets for nonneutralizing effector responses. In this study, we compared the antigenic properties of Env expressed in situ on pseudovirion virus-like particle (VLP) surfaces and soluble gp120 using harmonized enzyme-linked immunosorbent assays (ELISAs) and a panel of 51 monoclonal antibodies (MAbs). Only 32 of 46 soluble gp120-reactive MAbs recognized the primary UNC gp160 antigen of VLPs. Indeed, many epitopes were poorly exposed (C1, V2, C1-C4, C4, C4-V3, CD4 induced [CD4i], and PGT group 3) or obscured (C2, C5, and C1-C5) on VLPs. In further studies, VLP Env exhibited an increased degree of inter-MAb competition, the epicenter of which was the base of the V3 loop, where PGT, 2G12, V3, and CD4 binding site specificities competed. UNC gp160 also underwent more drastic soluble CD4 (sCD4)-induced conformational changes than soluble gp120, exposing CD4i, C1-C4, and V2 epitopes. A greater propensity of UNC gp160 to undergo conformational changes was also suggested by the induction of CD4i MAb binding to VLPs by a V3 MAb as well as by soluble CD4. The same effect was not observed for soluble gp120. Taken together, our data suggest that membrane-expressed UNC gp160 exists in a less "triggered" conformational state than soluble gp120 and that MAb binding to UNC gp160 tends to have greater conformational consequences.

Cleavage strongly influences whether soluble HIV-1 envelope glycoprotein trimers adopt a native-like conformation

“…For example, multiple non-NAbs are much more reactive with various gp120 epitopes on uncleaved trimers, as are ones to the six-helix bundle form of gp41 (13,(26)(27)(28). Thus, we believe that the aberrant soluble gp140 structures that we have visualized by EM will have their counterparts on the surface of cells expressing uncleaved Env proteins.…”

“…Cleaved and uncleaved, soluble or membrane-bound trimers express NAb and non-NAb epitopes very differently (13,(26)(27)(28). We therefore assessed the antigenicity of purified, D7324-tagged versions of SOSIP.R6, SOSIP.SEKS, and WT.SEKS gp140 trimers.…”

“…Thus, we believe that the aberrant soluble gp140 structures that we have visualized by EM will have their counterparts on the surface of cells expressing uncleaved Env proteins. Perhaps additional stability imparted by the transmembrane region will reduce the extent to which membrane-associated trimers decay into AC forms, but a wealth of antigenicity data strongly argues that such structures are commonplace on the surface of cells expressing uncleaved gp160s (13,(26)(27)(28).…”

Identification of Human Anti-HIV gp160 Monoclonal Antibodies That Make Effective Immunotoxins

“…In contrast, sCD4 (57) and MAbs directed against gp41 (58) do cause gp120/gp41 dissociation. These cooperative interactions probably result from the structural flexibility of this Env in its native state and the conformational alterations that have been observed to result upon CD4 or MAb binding (59)(60)(61).…”

HIV Envelope gp120 Alters T Cell Receptor Mobilization in the Immunological Synapse of Uninfected CD4 T Cells and Augments T Cell Activation

“…Past studies also have reported the activation of Lck and the downstream signals upon CD4 engagement by the HIV virions or the HIV Env expressed on cells (51)(52)(53). On the surfaces of virions and infected cells, heterogeneous species of the HIV Env glycoproteins are expressed (54)(55)(56), including nonfunctional Env proteins that cannot partake in the virus entry process but may have immunomodulatory activities demonstrated in this study.…”