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Association of VH4-59 Antibody Variable Gene Usage with Recognition of an Immunodominant Epitope on the HIV-1 Gag Protein.

Chukwuma VU, Hicar MD, Chen X, Nicholas KJ, Joyner A, Kalams SA, Landucci G, Forthal DN, Spearman PW, Crowe JE - PLoS ONE (2015)

Bottom Line: We found no evidence that mAb 3E4 exhibited any function in laboratory studies aimed at elucidating the immunologic activity, including assays for neutralization, Ab-dependent cell-mediated virus inhibition, or enhanced T cell reactivity caused by Gag-3E4 complexes.The findings suggest this immunodominant epitope in Gag protein, which is associated with VH4-59 germline gene usage, may induce a high level of B cells that encode binding but non-functional antibodies that occupy significant repertoire space following HIV infection.The studies define an additional specific molecular mechanism in the immune distraction activity of the HIV virion.

View Article: PubMed Central - PubMed

Affiliation: Department of Pathology, Microbiology and Immunology, Vanderbilt University, Nashville, Tennessee, United States of America.

ABSTRACT
The human antibody response against HIV-1 infection recognizes diverse antigenic subunits of the virion, and includes a high level of antibodies to the Gag protein. We report here the isolation and characterization of a subset of Gag-specific human monoclonal antibodies (mAbs) that were prevalent in the antibody repertoire of an HIV-infected individual. Several lineages of Gag-specifc mAbs were encoded by a single antibody heavy chain variable region, VH4-59, and a representative antibody from this group designated mAb 3E4 recognized a linear epitope on the globular head of the p17 subunit of Gag. We found no evidence that mAb 3E4 exhibited any function in laboratory studies aimed at elucidating the immunologic activity, including assays for neutralization, Ab-dependent cell-mediated virus inhibition, or enhanced T cell reactivity caused by Gag-3E4 complexes. The findings suggest this immunodominant epitope in Gag protein, which is associated with VH4-59 germline gene usage, may induce a high level of B cells that encode binding but non-functional antibodies that occupy significant repertoire space following HIV infection. The studies define an additional specific molecular mechanism in the immune distraction activity of the HIV virion.

No MeSH data available.


Related in: MedlinePlus

Generation of p17 Matrix deletion mutants.The full amino acid sequence alignment of the wild-type (WT) Matrix protein and the 10 Matrix deletion mutants. Conserved residues in the Matrix deletion mutants are indicated by the * symbol, and deleted resides indicated by the—symbol.
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pone.0133509.g005: Generation of p17 Matrix deletion mutants.The full amino acid sequence alignment of the wild-type (WT) Matrix protein and the 10 Matrix deletion mutants. Conserved residues in the Matrix deletion mutants are indicated by the * symbol, and deleted resides indicated by the—symbol.

Mentions: Next, we used a collection of overlapping peptides based on the HIV-1 group M consensus sequence of Gag to map the epitope of mAb 3E4. By testing binding to overlapping linear peptides, we determined that mAb 3E4 bound to a peptide in the p17 region of the Gag protein (Fig 4A). This peptide, with an amino acid sequence of HLVWASRELERFALN, is located on the surface of the Gag protein in the native structure (Fig 4B), exposing it for antigen-antibody interactions. Murine antibodies to this epitope have been described and have been shown to bind the surface of infected cells [28]. Given the alpha helical nature of the region where the peptide was located, we were interested in determining whether mutations in this region would affect the binding of mAb 3E4 to the p17 protein. We hypothesized that mutations in the globular head of the p17 protein would abrogate binding of mAb 3E4, due to a loss of stability associated with breaking alpha helices and the resulting conformational shifts that would occur. We generated ten p17 deletion mutants to test this hypothesis, by making overlapping amino acid deletions in the regions around the mAb 3E4 epitope on the p17 protein (Fig 5). We tested binding of mAb 3E4 to these p17 deletion mutants by western blot analysis, and observed a loss of binding to deletion mutants #1, #3, #4, #5, #6, and #7 shown in blue, yellow, magenta, cyan, orange or wheat colors, respectively (Fig 4C and 4D). These results showed that mutations in the globular head of the p17 protein abrogate the binding of mAb 3E4 to p17.


Association of VH4-59 Antibody Variable Gene Usage with Recognition of an Immunodominant Epitope on the HIV-1 Gag Protein.

Chukwuma VU, Hicar MD, Chen X, Nicholas KJ, Joyner A, Kalams SA, Landucci G, Forthal DN, Spearman PW, Crowe JE - PLoS ONE (2015)

Generation of p17 Matrix deletion mutants.The full amino acid sequence alignment of the wild-type (WT) Matrix protein and the 10 Matrix deletion mutants. Conserved residues in the Matrix deletion mutants are indicated by the * symbol, and deleted resides indicated by the—symbol.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4520566&req=5

pone.0133509.g005: Generation of p17 Matrix deletion mutants.The full amino acid sequence alignment of the wild-type (WT) Matrix protein and the 10 Matrix deletion mutants. Conserved residues in the Matrix deletion mutants are indicated by the * symbol, and deleted resides indicated by the—symbol.
Mentions: Next, we used a collection of overlapping peptides based on the HIV-1 group M consensus sequence of Gag to map the epitope of mAb 3E4. By testing binding to overlapping linear peptides, we determined that mAb 3E4 bound to a peptide in the p17 region of the Gag protein (Fig 4A). This peptide, with an amino acid sequence of HLVWASRELERFALN, is located on the surface of the Gag protein in the native structure (Fig 4B), exposing it for antigen-antibody interactions. Murine antibodies to this epitope have been described and have been shown to bind the surface of infected cells [28]. Given the alpha helical nature of the region where the peptide was located, we were interested in determining whether mutations in this region would affect the binding of mAb 3E4 to the p17 protein. We hypothesized that mutations in the globular head of the p17 protein would abrogate binding of mAb 3E4, due to a loss of stability associated with breaking alpha helices and the resulting conformational shifts that would occur. We generated ten p17 deletion mutants to test this hypothesis, by making overlapping amino acid deletions in the regions around the mAb 3E4 epitope on the p17 protein (Fig 5). We tested binding of mAb 3E4 to these p17 deletion mutants by western blot analysis, and observed a loss of binding to deletion mutants #1, #3, #4, #5, #6, and #7 shown in blue, yellow, magenta, cyan, orange or wheat colors, respectively (Fig 4C and 4D). These results showed that mutations in the globular head of the p17 protein abrogate the binding of mAb 3E4 to p17.

Bottom Line: We found no evidence that mAb 3E4 exhibited any function in laboratory studies aimed at elucidating the immunologic activity, including assays for neutralization, Ab-dependent cell-mediated virus inhibition, or enhanced T cell reactivity caused by Gag-3E4 complexes.The findings suggest this immunodominant epitope in Gag protein, which is associated with VH4-59 germline gene usage, may induce a high level of B cells that encode binding but non-functional antibodies that occupy significant repertoire space following HIV infection.The studies define an additional specific molecular mechanism in the immune distraction activity of the HIV virion.

View Article: PubMed Central - PubMed

Affiliation: Department of Pathology, Microbiology and Immunology, Vanderbilt University, Nashville, Tennessee, United States of America.

ABSTRACT
The human antibody response against HIV-1 infection recognizes diverse antigenic subunits of the virion, and includes a high level of antibodies to the Gag protein. We report here the isolation and characterization of a subset of Gag-specific human monoclonal antibodies (mAbs) that were prevalent in the antibody repertoire of an HIV-infected individual. Several lineages of Gag-specifc mAbs were encoded by a single antibody heavy chain variable region, VH4-59, and a representative antibody from this group designated mAb 3E4 recognized a linear epitope on the globular head of the p17 subunit of Gag. We found no evidence that mAb 3E4 exhibited any function in laboratory studies aimed at elucidating the immunologic activity, including assays for neutralization, Ab-dependent cell-mediated virus inhibition, or enhanced T cell reactivity caused by Gag-3E4 complexes. The findings suggest this immunodominant epitope in Gag protein, which is associated with VH4-59 germline gene usage, may induce a high level of B cells that encode binding but non-functional antibodies that occupy significant repertoire space following HIV infection. The studies define an additional specific molecular mechanism in the immune distraction activity of the HIV virion.

No MeSH data available.


Related in: MedlinePlus