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Differential binding of neutralizing and non-neutralizing antibodies to native-like soluble HIV-1 Env trimers, uncleaved Env proteins, and monomeric subunits.

Yasmeen A, Ringe R, Derking R, Cupo A, Julien JP, Burton DR, Ward AB, Wilson IA, Sanders RW, Moore JP, Klasse PJ - Retrovirology (2014)

Bottom Line: Trimerization and the proteolytic processing of Env precursors into gp120 and gp41 profoundly alter antigenicity, but soluble cleaved trimers are too unstable to serve as immunogens.We compared Fab and IgG versions of bNAbs and validated a bivalent model of IgG binding.High off-rate constants and low stoichiometric estimates of NAb binding were associated with large amounts of residual infectivity after NAb neutralization of the corresponding BG505.T332N pseudovirus.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Microbiology and Immunology, Weill Cornell Medical College, Cornell University, New York, USA. pek2003@med.cornell.edu.

ABSTRACT

Background: The trimeric envelope glycoproteins (Env) on the surface of HIV-1 virions are the targets for neutralizing antibodies (NAbs). No candidate HIV-1 immunogen has yet induced potent, broadly active NAbs (bNAbs). Part of the explanation may be that previously tested Env proteins inadequately mimic the functional, native Env complex. Trimerization and the proteolytic processing of Env precursors into gp120 and gp41 profoundly alter antigenicity, but soluble cleaved trimers are too unstable to serve as immunogens. By introducing stabilizing mutations (SOSIP), we constructed soluble, cleaved Env trimers derived from the HIV-1 subtype A isolate BG505 that resemble native Env spikes on virions both structurally and antigenically.

Results: We used surface plasmon resonance (SPR) to quantify antibody binding to different forms of BG505 Env: the proteolytically cleaved SOSIP.664 trimers, cleaved gp120-gp41ECTO protomers, and gp120 monomers. Non-NAbs to the CD4-binding site bound only marginally to the trimers but equally well to gp120-gp41ECTO protomers and gp120 monomers, whereas the bNAb VRC01, directed to the CD4bs, bound to all three forms. In contrast, bNAbs to V1V2 glycan-dependent epitopes bound preferentially (PG9 and PG16) or exclusively (PGT145) to trimers. We also explored the antigenic consequences of three different features of SOSIP.664 gp140 trimers: the engineered inter-subunit disulfide bond, the trimer-stabilizing I559P change in gp41ECTO, and proteolytic cleavage at the gp120-gp41ECTO junction. Each of these three features incrementally promoted native-like trimer antigenicity. We compared Fab and IgG versions of bNAbs and validated a bivalent model of IgG binding. The NAbs showed widely divergent binding kinetics and degrees of binding to native-like BG505 SOSIP.664. High off-rate constants and low stoichiometric estimates of NAb binding were associated with large amounts of residual infectivity after NAb neutralization of the corresponding BG505.T332N pseudovirus.

Conclusions: The antigenicity and structural integrity of cleaved BG505 SOSIP.664 trimers render these proteins good mimics of functional Env spikes on virions. In contrast, uncleaved gp140s antigenically resemble individual gp120-gp41ECTO protomers and gp120 monomers, but not native trimers. Although NAb binding to functional trimers may thus be both necessary and sufficient for neutralization, the kinetics and stoichiometry of the interaction influence the neutralizing efficacy of individual NAbs.

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The effects of cleavage and trimer-stabilizing mutations on Env antigenicity. The sensorgrams show the binding (RU) of the listed MAbs to BG505 SOSIP.664 trimers (labeled “SOSIP.R6”) and five mutated variants thereof (see Results). MAbs b12, b6, F240, 14e, and 19b do not neutralize the corresponding BG505.T332N virus, whereas VRC01, 2G12, PG16, and PGT145 all do. The SPR method was the same as for Figure 1, except that the MAbs were injected at 500 nM. The sensorgrams show one of two replicates.
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Figure 2: The effects of cleavage and trimer-stabilizing mutations on Env antigenicity. The sensorgrams show the binding (RU) of the listed MAbs to BG505 SOSIP.664 trimers (labeled “SOSIP.R6”) and five mutated variants thereof (see Results). MAbs b12, b6, F240, 14e, and 19b do not neutralize the corresponding BG505.T332N virus, whereas VRC01, 2G12, PG16, and PGT145 all do. The SPR method was the same as for Figure 1, except that the MAbs were injected at 500 nM. The sensorgrams show one of two replicates.

Mentions: We studied the binding of nine MAbs to six forms of Env: SOSIP.R6 (i.e., SOSIP.664) is the fully cleaved and stabilized form; WT.SEKS lacks both the SOSIP mutations and the cleavage site; SOSIP.SEKS lacks only the cleavage site; SOS.R6 lacks only the I559P mutation; SOS.SEKS lacks the I559P mutation and the cleavage site; IP.SEKS lacks the SOS link between gp120 and gp41ECTO as well as the cleavage site (Figure 2). Note that cleaved Env lacking the SOS linkage could not be studied because with that construct gp120 completely dissociates from gp41ECTO.


Differential binding of neutralizing and non-neutralizing antibodies to native-like soluble HIV-1 Env trimers, uncleaved Env proteins, and monomeric subunits.

Yasmeen A, Ringe R, Derking R, Cupo A, Julien JP, Burton DR, Ward AB, Wilson IA, Sanders RW, Moore JP, Klasse PJ - Retrovirology (2014)

The effects of cleavage and trimer-stabilizing mutations on Env antigenicity. The sensorgrams show the binding (RU) of the listed MAbs to BG505 SOSIP.664 trimers (labeled “SOSIP.R6”) and five mutated variants thereof (see Results). MAbs b12, b6, F240, 14e, and 19b do not neutralize the corresponding BG505.T332N virus, whereas VRC01, 2G12, PG16, and PGT145 all do. The SPR method was the same as for Figure 1, except that the MAbs were injected at 500 nM. The sensorgrams show one of two replicates.
© Copyright Policy - open-access
Related In: Results  -  Collection

License 1 - License 2
Show All Figures
getmorefigures.php?uid=PMC4067080&req=5

Figure 2: The effects of cleavage and trimer-stabilizing mutations on Env antigenicity. The sensorgrams show the binding (RU) of the listed MAbs to BG505 SOSIP.664 trimers (labeled “SOSIP.R6”) and five mutated variants thereof (see Results). MAbs b12, b6, F240, 14e, and 19b do not neutralize the corresponding BG505.T332N virus, whereas VRC01, 2G12, PG16, and PGT145 all do. The SPR method was the same as for Figure 1, except that the MAbs were injected at 500 nM. The sensorgrams show one of two replicates.
Mentions: We studied the binding of nine MAbs to six forms of Env: SOSIP.R6 (i.e., SOSIP.664) is the fully cleaved and stabilized form; WT.SEKS lacks both the SOSIP mutations and the cleavage site; SOSIP.SEKS lacks only the cleavage site; SOS.R6 lacks only the I559P mutation; SOS.SEKS lacks the I559P mutation and the cleavage site; IP.SEKS lacks the SOS link between gp120 and gp41ECTO as well as the cleavage site (Figure 2). Note that cleaved Env lacking the SOS linkage could not be studied because with that construct gp120 completely dissociates from gp41ECTO.

Bottom Line: Trimerization and the proteolytic processing of Env precursors into gp120 and gp41 profoundly alter antigenicity, but soluble cleaved trimers are too unstable to serve as immunogens.We compared Fab and IgG versions of bNAbs and validated a bivalent model of IgG binding.High off-rate constants and low stoichiometric estimates of NAb binding were associated with large amounts of residual infectivity after NAb neutralization of the corresponding BG505.T332N pseudovirus.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Microbiology and Immunology, Weill Cornell Medical College, Cornell University, New York, USA. pek2003@med.cornell.edu.

ABSTRACT

Background: The trimeric envelope glycoproteins (Env) on the surface of HIV-1 virions are the targets for neutralizing antibodies (NAbs). No candidate HIV-1 immunogen has yet induced potent, broadly active NAbs (bNAbs). Part of the explanation may be that previously tested Env proteins inadequately mimic the functional, native Env complex. Trimerization and the proteolytic processing of Env precursors into gp120 and gp41 profoundly alter antigenicity, but soluble cleaved trimers are too unstable to serve as immunogens. By introducing stabilizing mutations (SOSIP), we constructed soluble, cleaved Env trimers derived from the HIV-1 subtype A isolate BG505 that resemble native Env spikes on virions both structurally and antigenically.

Results: We used surface plasmon resonance (SPR) to quantify antibody binding to different forms of BG505 Env: the proteolytically cleaved SOSIP.664 trimers, cleaved gp120-gp41ECTO protomers, and gp120 monomers. Non-NAbs to the CD4-binding site bound only marginally to the trimers but equally well to gp120-gp41ECTO protomers and gp120 monomers, whereas the bNAb VRC01, directed to the CD4bs, bound to all three forms. In contrast, bNAbs to V1V2 glycan-dependent epitopes bound preferentially (PG9 and PG16) or exclusively (PGT145) to trimers. We also explored the antigenic consequences of three different features of SOSIP.664 gp140 trimers: the engineered inter-subunit disulfide bond, the trimer-stabilizing I559P change in gp41ECTO, and proteolytic cleavage at the gp120-gp41ECTO junction. Each of these three features incrementally promoted native-like trimer antigenicity. We compared Fab and IgG versions of bNAbs and validated a bivalent model of IgG binding. The NAbs showed widely divergent binding kinetics and degrees of binding to native-like BG505 SOSIP.664. High off-rate constants and low stoichiometric estimates of NAb binding were associated with large amounts of residual infectivity after NAb neutralization of the corresponding BG505.T332N pseudovirus.

Conclusions: The antigenicity and structural integrity of cleaved BG505 SOSIP.664 trimers render these proteins good mimics of functional Env spikes on virions. In contrast, uncleaved gp140s antigenically resemble individual gp120-gp41ECTO protomers and gp120 monomers, but not native trimers. Although NAb binding to functional trimers may thus be both necessary and sufficient for neutralization, the kinetics and stoichiometry of the interaction influence the neutralizing efficacy of individual NAbs.

Show MeSH
Related in: MedlinePlus