Structural characteristics correlate with immune responses induced by HIV envelope glycoprotein vaccines

Abstract

HIV envelope glycoprotein (Env) is the target for inducing neutralizing antibodies. Env is present on the virus surface as a trimer, and, upon binding to CD4, a cascade of events leads to structural rearrangement exposing the co-receptor binding site and entry into the CD4+ host target cells. We have designed monomeric and trimeric Env constructs with and without deletion of the variable loop 2 (ΔV2) from SF162, a subtype B primary isolate, and performed biophysical, biochemical and immunological studies to establish a potential structure–functional relationship. We expressed these Envs in CHO cells, purified the proteins to homogeneity and performed biophysical studies to define the binding properties to CD4, structural characteristics and exposure of epitopes recognized by b12 and CD4i mAb (17B) on both full-length and mutant HIV Env proteins. Parameters evaluated include oligomerization state, number and affinity of CD4 binding sites, enthalpy and entropy of the Env–CD4 interaction and affinity for b12 and 17b mAbs. We observed one CD4 binding site per monomer and three active CD4 binding sites per trimer. A40-fold difference in affinity of the gp120 monomer vs. the o-gp140 trimer towards CD4 was observed (Kd = 58 nM and 1.5 nM, respectively),whereas only a 2-fold difference was observed for the V2 deleted Envs (Kd of gp120ΔV2 = 19 nM, Kd of o-gp140DV2 = 9.3 nM). Monomers had 3-fold higher affinity to the mAb 17b and at least 3-fold weaker affinity to b12 compared to trimers, with gp120DV2 having the weakest affinity for b12 (Kd = 446 nM). Affinity of CD4 binding correlated with proportion of the antibodies induced against the conformational epitopes by the corresponding Envs, and changes in mAb binding correlated with the induction of antibodies directed against linear epitopes. Furthermore,biophysical analysis reveals that the V2 deletion has broad structural implications in the monomer not shared by the trimer, and these changes are reflected in the quality of the immune responses induced in rabbits. These data suggest that biophysical characteristics of HIV Env, such as affinity for CD4, and exposure of important neutralizing epitopes, such as those recognized by b12 mAb, may be important predictors of its in vivo efficacy and may serve as important surrogate markers for screening Env structures as potential vaccine candidates.

Quaternary structures of HIV Env immunogen exhibit conformational vicissitudes and interface diminution elicited by ligand binding

“…The gp120 subunit rotates, from having its CD4BS perpendicularly accessible prior to the binding, to a location laterally exposed from gp120, such that CD4 does not sterically occlude the coreceptor binding site, thus maximizing CCR5 or CXCR4 accessibility to the bridging sheet and V3 loop. Previous work has established that all three CD4 binding sites are exposed in the trimer construct (25), corroborating the exposed nature of the CD4BS as evidenced by our unliganded gp140 docking. However, the Liu model did not account for the observed tilt away from the z-axis, which flattens the trimer and exposes the threefold axis, the location where the N-terminal fusion peptide of gp41 would ostensibly protrude from and insert into the host membrane.…”

Comparable Antigenicity and Immunogenicity of Oligomeric Forms of a Novel, Acute HIV-1 Subtype C gp145 Envelope for Use in Preclinical and Clinical Vaccine Research

“…In addition, the results obtained with CD4i MAb 17b, with or without sCD4, are perplexing. Another group (81) has reported that CD4i antibodies typically do not bind the structurally intact envelope protein in the absence of CD4; however, MAb 17b has been reported to bind to some trimeric gp140 Env proteins (82,83). MAb 17b did bind all of the CO6980v0c22 species, and this binding was substantially increased in the presence of sCD4 (Table 1).…”

Comparative Immunogenicity of Subtype A Human Immunodeficiency Virus Type 1 Envelope Exhibiting Differential Exposure of Conserved Neutralization Epitopes

“…Next, the captured oligomeric gp140 protein was passed over DEAE and ceramic hydroxyapatite (CHAP) columns, allowing oligomeric gp140 to flow through both columns and contaminating proteins to bind to one column or the other. All the column fractions were analyzed on SDS-PAGE and also in a CD4 receptorbinding assay as described elsewhere in detail (58)(59)(60).…”