The rhesus DH3-15*01 exhibits structural plasticity and encodes a unique anionic motif. (a) HCDR3 structures from rhesus PGT145-like Fab’s in complex with envelope trimers. DH3-15*01 conserved five-residue motif (EDDYG) positions are colored and labeled in dark red, and the remaining D gene positions are colored and labeled in pink. Conserved motif position labels are italicized when subjected to SHM. D gene position side chains are shown in ball-and-stick representation with nitrogen atoms colored blue, oxygen atoms colored red, and sulfur atoms colored yellow (Kabat numbering). The remaining HCDR3 residues are colored gray with side chains hidden. Below each structure is an alignment of the germline DH3-15*01 coding fragment that was acquired during VDJ recombination (bottom) with the sequence at these positions in the mature antibody (top; Kabat numbering). Somatic mutations that conserve side chain aromaticity or anionic charge are depicted in bold, while discordant somatic mutations are underlined. Sites of tyrosine sulfation are highlighted in green. The functional role of the conserved five-residue motif segment is written above the alignment. (b) HCDR3 structures from rhesus PG9-like Fab’s in complex with envelope trimers. Structures are depicted similarly to panel a. (c) HCDR3 structures from rhesus VRC26-like Fab’s in complex with SOSIP trimers. Structures are depicted similarly to panel a. Residue insertions in these structures and sequence alignments are colored blue. (d) Proportion of reading frame usage among naïve rhesus B cells derived from DH3 genes in the peripheral Indian rhesus macaque repertoire. DH3-15*01 is highlighted in red here and in the remaining panels. B cells derived from rhesus DH3-19*01 are exceedingly rare and therefore excluded from analysis. (e) Frequency of DH3 family usage in reading frame two among all naïve B cells in the peripheral rhesus repertoire. (f) HCDR3 length distributions of naïve rhesus B cells in the peripheral rhesus repertoire derived from DH3 genes in reading frame two. (g) HCDR3 net charge distributions of naïve rhesus B cells in the peripheral repertoire derived from DH3 genes in reading frame two. The net charge of DH3-15*01–derived HCDR3s is more anionic (****, P < 0.00001 Student’s t test) than all other groups. Charge calculations only consider amino acid residues and not predicted sites of tyrosine sulfation.
Figure 6.

The rhesus DH3-15*01 exhibits structural plasticity and encodes a unique anionic motif. (a) HCDR3 structures from rhesus PGT145-like Fab’s in complex with envelope trimers. DH3-15*01 conserved five-residue motif (EDDYG) positions are colored and labeled in dark red, and the remaining D gene positions are colored and labeled in pink. Conserved motif position labels are italicized when subjected to SHM. D gene position side chains are shown in ball-and-stick representation with nitrogen atoms colored blue, oxygen atoms colored red, and sulfur atoms colored yellow (Kabat numbering). The remaining HCDR3 residues are colored gray with side chains hidden. Below each structure is an alignment of the germline DH3-15*01 coding fragment that was acquired during VDJ recombination (bottom) with the sequence at these positions in the mature antibody (top; Kabat numbering). Somatic mutations that conserve side chain aromaticity or anionic charge are depicted in bold, while discordant somatic mutations are underlined. Sites of tyrosine sulfation are highlighted in green. The functional role of the conserved five-residue motif segment is written above the alignment. (b) HCDR3 structures from rhesus PG9-like Fab’s in complex with envelope trimers. Structures are depicted similarly to panel a. (c) HCDR3 structures from rhesus VRC26-like Fab’s in complex with SOSIP trimers. Structures are depicted similarly to panel a. Residue insertions in these structures and sequence alignments are colored blue. (d) Proportion of reading frame usage among naïve rhesus B cells derived from DH3 genes in the peripheral Indian rhesus macaque repertoire. DH3-15*01 is highlighted in red here and in the remaining panels. B cells derived from rhesus DH3-19*01 are exceedingly rare and therefore excluded from analysis. (e) Frequency of DH3 family usage in reading frame two among all naïve B cells in the peripheral rhesus repertoire. (f) HCDR3 length distributions of naïve rhesus B cells in the peripheral rhesus repertoire derived from DH3 genes in reading frame two. (g) HCDR3 net charge distributions of naïve rhesus B cells in the peripheral repertoire derived from DH3 genes in reading frame two. The net charge of DH3-15*01–derived HCDR3s is more anionic (****, P < 0.00001 Student’s t test) than all other groups. Charge calculations only consider amino acid residues and not predicted sites of tyrosine sulfation.

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