Supplementary Figure 1: Evolution of a second-generation H-2Ld scaffold.
(a) The protein architecture of the single chain m31r-CP template used for error prone PCR. (b) The enrichment of gain-of-function mutations in yeast scaffolds, during rounds of selections using TCR-tetramers. (c) The gain-of-function mutations that provided the brightest 2C-tetramer and 42F3-tetramer staining on the clonal yeast population m31r-CP-E3. Inset is the zoomed in depiction of the peptide linker-MHC-junctions and proximal selected mutations (sticks). (d) Mutations in the m31s-W167A used to display the C-terminally linked peptide libraries. (e) Staining of 96 clonal yeast populations isolated from the final round of selection for the 42F3 TCR. (f) The unique peptide sequences selected by 42F3 TCR observed in the sequencing sample. The P2 and P9 positions with restricted diversity are highlighted in black and blue respectively, and positions fully diversified are highlighted in red.
