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Viral escape from neutralizing antibodies in early subtype A HIV-1 infection drives an increase in autologous neutralization breadth.

Murphy MK, Yue L, Pan R, Boliar S, Sethi A, Tian J, Pfafferot K, Karita E, Allen SA, Cormier E, Goepfert PA, Borrow P, Robinson JE, Gnanakaran S, Hunter E, Kong XP, Derdeyn CA - PLoS Pathog. (2013)

Bottom Line: Crystal structures of the antigen-binding fragments (Fabs) revealed flat epitope contact surfaces, where minimal light chain mutation in 19.3H-L3 allowed for additional antigenic interactions.Our data demonstrate that this subject's first recognized nAb epitope elicited strain-specific mAbs, which incrementally acquired autologous breadth, and directed later B cell responses to target distinct portions of Env.This immune re-focusing could have triggered the evolution of cross-clade antibodies and suggests that exposure to a specific sequence of immune escape variants might promote broad humoral responses during HIV-1 infection.

View Article: PubMed Central - PubMed

Affiliation: Immunology and Molecular Pathogenesis Graduate Program, Emory University, Atlanta, Georgia, United States of America.

ABSTRACT
Antibodies that neutralize (nAbs) genetically diverse HIV-1 strains have been recovered from a subset of HIV-1 infected subjects during chronic infection. Exact mechanisms that expand the otherwise narrow neutralization capacity observed during early infection are, however, currently undefined. Here we characterized the earliest nAb responses in a subtype A HIV-1 infected Rwandan seroconverter who later developed moderate cross-clade nAb breadth, using (i) envelope (Env) glycoproteins from the transmitted/founder virus and twenty longitudinal nAb escape variants, (ii) longitudinal autologous plasma, and (iii) autologous monoclonal antibodies (mAbs). Initially, nAbs targeted a single region of gp120, which flanked the V3 domain and involved the alpha2 helix. A single amino acid change at one of three positions in this region conferred early escape. One immunoglobulin heavy chain and two light chains recovered from autologous B cells comprised two mAbs, 19.3H-L1 and 19.3H-L3, which neutralized the founder Env along with one or three of the early escape variants carrying these mutations, respectively. Neither mAb neutralized later nAb escape or heterologous Envs. Crystal structures of the antigen-binding fragments (Fabs) revealed flat epitope contact surfaces, where minimal light chain mutation in 19.3H-L3 allowed for additional antigenic interactions. Resistance to mAb neutralization arose in later Envs through alteration of two glycans spatially adjacent to the initial escape signatures. The cross-neutralizing nAbs that ultimately developed failed to target any of the defined V3-proximal changes generated during the first year of infection in this subject. Our data demonstrate that this subject's first recognized nAb epitope elicited strain-specific mAbs, which incrementally acquired autologous breadth, and directed later B cell responses to target distinct portions of Env. This immune re-focusing could have triggered the evolution of cross-clade antibodies and suggests that exposure to a specific sequence of immune escape variants might promote broad humoral responses during HIV-1 infection.

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Crystal structures of R880F mAbs 19.3H-L1 and 19.3H-L3.(A) CDR loops. A top view looking down at the antigen-binding site of 19.3H-L1 represented by ribbons. The framework regions of the light chain and heavy chain are colored cyan and green, respectively, while each CDR loop is colored separately. The side chains of the three VL CDR1 residues different between 19.3H-L1 and 19.3H-L3 are displayed. (B) ODA analysis of the Fab 19.3H-L1. The size/redness of each sphere is proportional to the binding strength of the region indicated. Note that the antigen-binding site is centered at VL CDR1 and VH CDR3. (C) The electrostatic surface potentials of the antigen-binding site of 19.3H-L1. Red and blue coloration represents the negatively and positively charged regions, respectively, while a dashed line encircles the flat surface of the antigen-binding site. (D) The three VL CDR1 amino acid differences, S27T, G29T, and Y32F, between 19.3H-L1 (cyan) and 19.3H-L3 (yellow).
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ppat-1003173-g007: Crystal structures of R880F mAbs 19.3H-L1 and 19.3H-L3.(A) CDR loops. A top view looking down at the antigen-binding site of 19.3H-L1 represented by ribbons. The framework regions of the light chain and heavy chain are colored cyan and green, respectively, while each CDR loop is colored separately. The side chains of the three VL CDR1 residues different between 19.3H-L1 and 19.3H-L3 are displayed. (B) ODA analysis of the Fab 19.3H-L1. The size/redness of each sphere is proportional to the binding strength of the region indicated. Note that the antigen-binding site is centered at VL CDR1 and VH CDR3. (C) The electrostatic surface potentials of the antigen-binding site of 19.3H-L1. Red and blue coloration represents the negatively and positively charged regions, respectively, while a dashed line encircles the flat surface of the antigen-binding site. (D) The three VL CDR1 amino acid differences, S27T, G29T, and Y32F, between 19.3H-L1 (cyan) and 19.3H-L3 (yellow).

Mentions: To interrogate the antigen-binding site characteristics of R880F mAbs that influenced their distinct neutralization profiles, crystal structures of the 19.3H-L1 and 19.3H-L3 Fabs were determined to the resolutions of 1.7 Å (Figure 7A) and 2.7 Å, respectively (Table S1). Although the two Fabs were crystallized in different space groups, the resultant structures were highly similar, with root mean square deviations less than 1 Å when all of the Cα atoms were superimposed (data not shown). Several structural analyses were employed, including calculations of Optical Docking Area (ODA, shown in Figure 7B, which predicted the antigen-binding sites by calculating the desolvation free energy of the surfaces), surface pockets, and electrostatic surface potentials. ODA analyses indicated that the antigen-binding sites of 19.3H-L1 and 19.3H-L3 were very flat, forming roughly rectangular shapes approximately 15 Å wide and 30 Å long on top of the six CDR loops (Figure 7C). No pockets existed in these binding surfaces, and the shared CDR H3, although it was 18 amino acids long (Kabat numbering scheme [33]), did not protrude. Such flat antigen-binding sites likely interact with epitopes formed by residues also on planar surfaces (i.e. flat-surface antigen-antibody contacts). Electrostatic surface potential analyses showed that the 19.3H-L1 and 19.3H-L3 antigen-binding sites were essentially neutral; a couple of slightly positive regions along one side of the rectangular contact area counterbalanced a slightly negative opposite region (Figure 7C, blue and red patches, respectively).


Viral escape from neutralizing antibodies in early subtype A HIV-1 infection drives an increase in autologous neutralization breadth.

Murphy MK, Yue L, Pan R, Boliar S, Sethi A, Tian J, Pfafferot K, Karita E, Allen SA, Cormier E, Goepfert PA, Borrow P, Robinson JE, Gnanakaran S, Hunter E, Kong XP, Derdeyn CA - PLoS Pathog. (2013)

Crystal structures of R880F mAbs 19.3H-L1 and 19.3H-L3.(A) CDR loops. A top view looking down at the antigen-binding site of 19.3H-L1 represented by ribbons. The framework regions of the light chain and heavy chain are colored cyan and green, respectively, while each CDR loop is colored separately. The side chains of the three VL CDR1 residues different between 19.3H-L1 and 19.3H-L3 are displayed. (B) ODA analysis of the Fab 19.3H-L1. The size/redness of each sphere is proportional to the binding strength of the region indicated. Note that the antigen-binding site is centered at VL CDR1 and VH CDR3. (C) The electrostatic surface potentials of the antigen-binding site of 19.3H-L1. Red and blue coloration represents the negatively and positively charged regions, respectively, while a dashed line encircles the flat surface of the antigen-binding site. (D) The three VL CDR1 amino acid differences, S27T, G29T, and Y32F, between 19.3H-L1 (cyan) and 19.3H-L3 (yellow).
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ppat-1003173-g007: Crystal structures of R880F mAbs 19.3H-L1 and 19.3H-L3.(A) CDR loops. A top view looking down at the antigen-binding site of 19.3H-L1 represented by ribbons. The framework regions of the light chain and heavy chain are colored cyan and green, respectively, while each CDR loop is colored separately. The side chains of the three VL CDR1 residues different between 19.3H-L1 and 19.3H-L3 are displayed. (B) ODA analysis of the Fab 19.3H-L1. The size/redness of each sphere is proportional to the binding strength of the region indicated. Note that the antigen-binding site is centered at VL CDR1 and VH CDR3. (C) The electrostatic surface potentials of the antigen-binding site of 19.3H-L1. Red and blue coloration represents the negatively and positively charged regions, respectively, while a dashed line encircles the flat surface of the antigen-binding site. (D) The three VL CDR1 amino acid differences, S27T, G29T, and Y32F, between 19.3H-L1 (cyan) and 19.3H-L3 (yellow).
Mentions: To interrogate the antigen-binding site characteristics of R880F mAbs that influenced their distinct neutralization profiles, crystal structures of the 19.3H-L1 and 19.3H-L3 Fabs were determined to the resolutions of 1.7 Å (Figure 7A) and 2.7 Å, respectively (Table S1). Although the two Fabs were crystallized in different space groups, the resultant structures were highly similar, with root mean square deviations less than 1 Å when all of the Cα atoms were superimposed (data not shown). Several structural analyses were employed, including calculations of Optical Docking Area (ODA, shown in Figure 7B, which predicted the antigen-binding sites by calculating the desolvation free energy of the surfaces), surface pockets, and electrostatic surface potentials. ODA analyses indicated that the antigen-binding sites of 19.3H-L1 and 19.3H-L3 were very flat, forming roughly rectangular shapes approximately 15 Å wide and 30 Å long on top of the six CDR loops (Figure 7C). No pockets existed in these binding surfaces, and the shared CDR H3, although it was 18 amino acids long (Kabat numbering scheme [33]), did not protrude. Such flat antigen-binding sites likely interact with epitopes formed by residues also on planar surfaces (i.e. flat-surface antigen-antibody contacts). Electrostatic surface potential analyses showed that the 19.3H-L1 and 19.3H-L3 antigen-binding sites were essentially neutral; a couple of slightly positive regions along one side of the rectangular contact area counterbalanced a slightly negative opposite region (Figure 7C, blue and red patches, respectively).

Bottom Line: Crystal structures of the antigen-binding fragments (Fabs) revealed flat epitope contact surfaces, where minimal light chain mutation in 19.3H-L3 allowed for additional antigenic interactions.Our data demonstrate that this subject's first recognized nAb epitope elicited strain-specific mAbs, which incrementally acquired autologous breadth, and directed later B cell responses to target distinct portions of Env.This immune re-focusing could have triggered the evolution of cross-clade antibodies and suggests that exposure to a specific sequence of immune escape variants might promote broad humoral responses during HIV-1 infection.

View Article: PubMed Central - PubMed

Affiliation: Immunology and Molecular Pathogenesis Graduate Program, Emory University, Atlanta, Georgia, United States of America.

ABSTRACT
Antibodies that neutralize (nAbs) genetically diverse HIV-1 strains have been recovered from a subset of HIV-1 infected subjects during chronic infection. Exact mechanisms that expand the otherwise narrow neutralization capacity observed during early infection are, however, currently undefined. Here we characterized the earliest nAb responses in a subtype A HIV-1 infected Rwandan seroconverter who later developed moderate cross-clade nAb breadth, using (i) envelope (Env) glycoproteins from the transmitted/founder virus and twenty longitudinal nAb escape variants, (ii) longitudinal autologous plasma, and (iii) autologous monoclonal antibodies (mAbs). Initially, nAbs targeted a single region of gp120, which flanked the V3 domain and involved the alpha2 helix. A single amino acid change at one of three positions in this region conferred early escape. One immunoglobulin heavy chain and two light chains recovered from autologous B cells comprised two mAbs, 19.3H-L1 and 19.3H-L3, which neutralized the founder Env along with one or three of the early escape variants carrying these mutations, respectively. Neither mAb neutralized later nAb escape or heterologous Envs. Crystal structures of the antigen-binding fragments (Fabs) revealed flat epitope contact surfaces, where minimal light chain mutation in 19.3H-L3 allowed for additional antigenic interactions. Resistance to mAb neutralization arose in later Envs through alteration of two glycans spatially adjacent to the initial escape signatures. The cross-neutralizing nAbs that ultimately developed failed to target any of the defined V3-proximal changes generated during the first year of infection in this subject. Our data demonstrate that this subject's first recognized nAb epitope elicited strain-specific mAbs, which incrementally acquired autologous breadth, and directed later B cell responses to target distinct portions of Env. This immune re-focusing could have triggered the evolution of cross-clade antibodies and suggests that exposure to a specific sequence of immune escape variants might promote broad humoral responses during HIV-1 infection.

Show MeSH
Related in: MedlinePlus