Intraprotomer masking of third variable loop (V3) epitopes by the first and second variable loops (V1V2) within the native HIV-1 envelope glycoprotein trimer

ABSTRACT We have described an oligomeric gp140 envelope glycoprotein from human immunodeficiency virus type 1 that is stabilized by an intermolecular disulfide bond between gp120 and the gp41 ectodomain, termed SOS gp140 (J. M. Binley, R. W. Sanders, B. Clas, N. Schuelke, A. Master, Y. Guo, F. Kajumo, D. J. Anselma, P. J. Maddon, W. C. Olson, and J. P. Moore, J. Virol. 74:627–643, 2000). In this protein, the protease cleavage site between gp120 and gp41 is fully utilized. Here we report the characterization of gp140 variants that have deletions in the first, second, and/or third variable loop (V1, V2, and V3 loops). The SOS disulfide bond formed efficiently in gp140s containing a single loop deletion or a combination deletion of the V1 and V2 loops. However, deletion of all three variable loops prevented formation of the SOS disulfide bond. Some variable-loop-deleted gp140s were not fully processed to their gp120 and gp41 constituents even when the furin protease was cotransfected. The exposure of the gp120-gp41 cleavage site is probably affected in these proteins, even though the disabling change is in a region of gp120 distal from the cleavage site. Antigenic characterization of the variable-loop-deleted SOS gp140 proteins revealed that deletion of the variable loops uncovers cryptic, conserved neutralization epitopes near the coreceptor-binding site on gp120. These modified, disulfide-stabilized glycoproteins might be useful as immunogens.

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Identification of Conserved and Variable Structures in the Human Immunodeficiency Virus gp120 Glycoprotein of Importance for CXCR4 Binding

Journal of Virology ◽

10.1128/jvi.76.21.10791-10800.2002 ◽

2002 ◽

Vol 76 (21) ◽

pp. 10791-10800 ◽

Cited By ~ 64

Author(s):

Stéphane Basmaciogullari ◽

Gregory J. Babcock ◽

Donald Van Ryk ◽

Woj Wojtowicz ◽

Joseph Sodroski

Keyword(s):

Human Immunodeficiency Virus ◽

Envelope Glycoprotein ◽

Chemokine Receptors ◽

Specific Binding ◽

Viral Glycoprotein ◽

Independent Manner ◽

Immunodeficiency Virus ◽

Variable Loops ◽

Gp120 Glycoprotein ◽

Hiv 1

ABSTRACT CD4 and the chemokine receptors, CXCR4 and CCR5, serve as receptors for human immunodeficiency virus type 1 (HIV-1). Binding of the HIV-1 gp120 envelope glycoprotein to the chemokine receptors normally requires prior interaction with CD4. Mapping the determinants on gp120 for the low-affinity interaction with CXCR4 has been difficult due to the nonspecific binding of this viral glycoprotein to cell surfaces. Here we examine the binding of a panel of gp120 mutants to paramagnetic proteoliposomes displaying CXCR4 on their surfaces. We show that the gp120 β19 strand and third variable (V3) loop contain residues important for CXCR4 interaction. Basic residues from both elements, as well as a conserved hydrophobic residue at the V3 tip, contribute to CXCR4 binding. Removal of the gp120 V1/V2 variable loops allows the envelope glycoprotein to bind CXCR4 in a CD4-independent manner. These results indicate that although some variable gp120 residues contribute to the specific binding to CCR5 or CXCR4, gp120 elements common to CXCR4- or CCR5-using strains are involved in the interaction with both coreceptors.

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The V4 and V5 Variable Loops of HIV-1 Envelope Glycoprotein Are Tolerant to Insertion of Green Fluorescent Protein and Are Useful Targets for Labeling

Journal of Biological Chemistry ◽

10.1074/jbc.m114.628610 ◽

2015 ◽

Vol 290 (24) ◽

pp. 15279-15291 ◽

Cited By ~ 17

Author(s):

Shuhei Nakane ◽

Aikichi Iwamoto ◽

Zene Matsuda

Keyword(s):

Green Fluorescent Protein ◽

Envelope Glycoprotein ◽

Fluorescent Protein ◽

Variable Loops ◽

Green Fluorescent ◽

Hiv 1

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HIV-1 Envelope Glycoprotein Variable Loops Are Indispensable for Envelope Structural Integrity and Virus Entry

PLoS ONE ◽

10.1371/journal.pone.0069789 ◽

2013 ◽

Vol 8 (8) ◽

pp. e69789 ◽

Cited By ~ 14

Author(s):

Tingting Yuan ◽

Jingjing Li ◽

Mei-Yun Zhang

Keyword(s):

Envelope Glycoprotein ◽

Structural Integrity ◽

Virus Entry ◽

Variable Loops ◽

Hiv 1

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Importance of the V1/V2 Loop Region of Simian-Human Immunodeficiency Virus Envelope Glycoprotein gp120 in Determining the Strain Specificity of the Neutralizing Antibody Response

Journal of Virology ◽

10.1128/jvi.01341-08 ◽

2008 ◽

Vol 82 (22) ◽

pp. 11054-11065 ◽

Cited By ~ 20

Author(s):

Melissa E. Laird ◽

Tatsuhiko Igarashi ◽

Malcolm A. Martin ◽

Ronald C. Desrosiers

Keyword(s):

Human Immunodeficiency Virus ◽

Envelope Glycoprotein ◽

Neutralizing Antibody ◽

Plasma Samples ◽

Strain Specificity ◽

Neutralizing Activity ◽

Immunodeficiency Virus ◽

Glycoprotein Gp120 ◽

Variable Loops ◽

Hiv 1

ABSTRACT Plasma samples from individuals infected with human immunodeficiency virus type 1 (HIV-1) are known to be highly strain specific in their ability to neutralize HIV-1 infectivity. Such plasma samples exhibit significant neutralizing activity against autologous HIV-1 isolates but typically exhibit little or no activity against heterologous strains, although some cross-neutralizing activity can develop late in infection. Monkeys infected with the simian-human immunodeficiency virus (SHIV) clone DH12 generated antibodies that neutralized SHIV DH12, but not SHIV KB9. Conversely, antibodies from monkeys infected with the SHIV clone KB9 neutralized SHIV KB9, but not SHIV DH12. To investigate the role of the variable loops of the HIV-1 envelope glycoprotein gp120 in determining this strain specificity, variable loops 1 and 2 (V1/V2), V3, or V4 were exchanged individually or in combination between SHIV DH12 and SHIV KB9. Despite the fact that both parental viruses exhibited significant infectivity and good replication in the cell lines examined, 3 of the 10 variable-loop chimeras exhibited such poor infectivity that they could not be used further for neutralization assays. These results indicate that a variable loop that is functional in the context of one particular envelope background will not necessarily function within another. The remaining seven replication-competent chimeras allowed unambiguous assignment of the sequences principally responsible for the strain specificity of the neutralizing activity present in SHIV-positive plasma. Exchange of the V1/V2 loop sequences conferred a dominant loss of sensitivity to neutralization by autologous plasma and a gain of sensitivity to neutralization by heterologous plasma. Substitution of V3 or V4 had little or no effect on the sensitivity to neutralization. These data demonstrate that the V1/V2 region of HIV-1 gp120 is principally responsible for the strain specificity of the neutralizing antibody response in monkeys infected with these prototypic SHIVs.

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Infectivity and Neutralization of Simian Immunodeficiency Virus with FLAG Epitope Insertion in gp120 Variable Loops

Journal of Virology ◽

10.1128/jvi.00831-07 ◽

2007 ◽

Vol 81 (20) ◽

pp. 10838-10848 ◽

Cited By ~ 15

Author(s):

Melissa E. Laird ◽

Ronald C. Desrosiers

Keyword(s):

Simian Immunodeficiency Virus ◽

Envelope Glycoprotein ◽

Flag Epitope ◽

Epitope Tag ◽

Recombinant Viruses ◽

Variable Loop ◽

Immunodeficiency Virus ◽

Variable Loops ◽

Glycoprotein Structure ◽

Global Increase

ABSTRACT A FLAG epitope tag was substituted within variable loop 1 (V1), 2 (V2), or 4 (V4) of the gp120 envelope glycoprotein of simian immunodeficiency virus strain 239 (SIV239) to evaluate the extent to which each variable loop may serve as a target for antibody-mediated neutralization. Two sites within each variable loop of SIV239 were chosen for individual epitope tag insertions. FLAG epitope substitutions were also made in the V1, V2, and V4 loops of a neutralization-sensitive derivative of SIV239, SIV316. Of the 10 FLAG-tagged recombinant viruses analyzed, three (SIV239FV1b, SIV239FV2b, and SIV239FV4a) replicated with kinetics similar to those of the parental strain, SIV239, in both CEMx174 cells and the immortalized rhesus monkey T-cell line 221. The SIV316FV1b and SIV316FV4a FLAG variants replicated with a substantial lag, and the five remaining recombinants did not replicate detectably. Both gp160 and gp120 from replication-competent FLAG variants could be immunoprecipitated from transfected 293T cells by the anti-gp120 rhesus monoclonal antibody (RhMAb) 3.11H, the anti-FLAG MAb M2, and CD4-immunoglobulin, whereas only unprocessed gp160 was detected in 293T cells transfected with replication-defective variants. Furthermore, gp120 was detectably incorporated only into virions that were infectious. SIV239FV1b was sensitive to neutralization by MAb M2, with a 50% inhibitory concentration of 1 μg/ml. Neither SIV239FV2b nor SIV239FV4a was sensitive to M2 neutralization. The ability of the M2 antibody to neutralize SIV239FV1b infectivity was associated with an increased ability of the M2 antibody to detect native, oligomeric SIV239FV1b envelope protein on the surfaces of cells relative to that for the other SIV FLAG variants. Furthermore, SIV239FV1b was globally more sensitive to antibody-mediated neutralization than was parental SIV239 when these strains were screened with a panel of anti-SIV MAbs of various specificities. These results indicate that the V1 loop can serve as an effective target for neutralization on SIV239FV1b. However, antibody-mediated neutralization of this variant, similar to that of other SIV239 variants that have been studied previously, was associated with a global increase in neutralization sensitivity. These results suggest that the variable loops on the neutralization-resistant SIV239 strain are difficult for antibodies to access effectively and that mutations that allow neutralization have global effects on the trimeric envelope glycoprotein structure and accessibility.

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