Figure 2. Substitution distributions of selected residues characterize residue interactions with antibody.

The enrichment for each particular substitution is reported for all substitutions arising from a single nucleotide substitution. (A) Epitope residues show enrichment of all or near all substitutions arising from a single nucleotide substitution consistent with the sidechain of these residues directly contacting paratope residues in the antibody (B) Some epitope residues show tolerance of particular substitution, demonstrated by the absence of these substitutions in the sequences obtained from the sorted populations, as a result of physicochemical similarity of the sidechains (C) Positions were only one or a few substitutions show statistically significant enrichment can be the result of particular substitutions that introduce negative interactions (D) Mutations to proline or glycine at many residues were ablative likely by preventing alpha helical secondary structure to develop. Mutations to residues critical to protein folding also led to reduction in affinity.