Human Anti-HIV-1 Env Recombinant Antibody (clone PGT124) (CAT#: PABL-137)

Figure 1 Mapping overlapping antibody epitope footprints

Pairwise alanine mutants were created for N137, N156, N301, and N332 glycans and the 324GDIRQAH330 residues of the V3 loop. Virus mutants were then tested for neutralization by PGT121, PGT124, PGT128, PGT130, PGDM12, PGDM21 and, as a control, the CD4 binding site antibody 12A12.

Sok, D., Pauthner, M., Briney, B., Lee, J. H., Saye-Francisco, K. L., Hsueh, J.,... & Bastidas, R. (2016). A prominent site of antibody vulnerability on HIV envelope incorporates a motif associated with CCR5 binding and its camouflaging glycans.Immunity, 45(1), 31-45.

Figure 2 Epitope recognition on native Env by V3-glycan antibodies

(A) Negative-stain EM of antibody Fabs on BG505 SOSIP.664. X-ray structures of gp120 liganded with PGT124 (PDB 4R2G), PGT122 (PDB 4TVP) and PGT128 (PDB 4TYG) were superimposed onto gp120 in the unliganded trimer volume using the 'MatchMaker' function in Chimera. All glycans shown are part of the 4TVP X–ray structure. For PGDM14 and PGDM21, the relative orientations of both Fabs were mapped onto a low resolution surface volume generated from the BG505 SOSIP.664 X-ray structure (PDB 4TVP). The reconstructions of PGDM14 (EMD-8181) and PGDM21 (EMD-8182) were superimposed onto this trimer volume using the 'Fit in Map' function in Chimera (https://www.cgl.ucsf.edu/chimera/). (B) Summary table of all GDIR-glycan bnAb families. Donor ID refers to the IAVI Protocol G donor ID (Simek et al., 2009), ND indicates not determined. (C) Proposed recognition of overlapping epitopes between V3-glycan antibodies and CD4i antibodies. In addition to making contacts to the 324GDIRQAH330 peptide region, CD4i antibodies require formation of the bridging sheet on gp120 following CD4 engagement in order to bind and therefore can only bind open conformations of the Env trimer. V3-glycan antibodies are capable of binding the immunogenic 324GDIRQAH330 peptide region on native Env in the absence of CD4 by penetrating the glycan shield and also directly binding the surrounding glycans.

Sok, D., Pauthner, M., Briney, B., Lee, J. H., Saye-Francisco, K. L., Hsueh, J.,... & Bastidas, R. (2016). A prominent site of antibody vulnerability on HIV envelope incorporates a motif associated with CCR5 binding and its camouflaging glycans.Immunity, 45(1), 31-45.

Figure 3 Key contacts to residues 324GDIRQAH330 on gp120 by GDIR-bnAbs and CD4i antibodies. Overlaid liganded structures of PGT122, PGT124 and PGT128 (shades of blue) with key contacts to the 324GDIRQAH330 residues on gp120 highlighted in red. These structures are overlaid with liganded structures of CD4i antibodies 17b, X5, and 412d (shades of yellow) with key contacts to the 324GDIRQAH330 residues on gp120 highlighted in green. Arg327 on gp120 for all structures is shown as sticks. All structures are aligned on gp120. (D) High-mannose patch antibodies were tested for competition with sCD4 on isolate JR-FL E168K/N192A Env displayed on the surface of 293T cells. The CD4 binding site antibody 12A12 was included as a positive control and the V3-specific antibody F425 was included as a negative control.

Sok, D., Pauthner, M., Briney, B., Lee, J. H., Saye-Francisco, K. L., Hsueh, J.,... & Bastidas, R. (2016). A prominent site of antibody vulnerability on HIV envelope incorporates a motif associated with CCR5 binding and its camouflaging glycans.Immunity, 45(1), 31-45.