Design of Protein Multi-specificity Using an Independent Sequence Search Reduces the Barrier to Low Energy Sequences
Fig 5
Recapitulation of evolutionary sequence profiles by multi-specificity design.
A. For each protein in the benchmark set, an evolutionary sequence profile (top) was calculated and compared to the sequences generated by MSD (bottom). A similarity score was calculated for each position and averaged over designed positions to measure how well design searches biologically relevant sequence space. Highlighted are example positions where designed sequences either agreed (blue) or disagreed (red) with naturally occurring sequences. The figure displays the designed amino acid profile for a subset of positions in the VH5-51 benchmark set. See methods for details on percent similarity calculation. Amino acids are colored according to chemical properties. B. RECON-generated designs were more similar to observed evolutionary sequence profiles than those produced by MPI_MSD. Percent similarity was averaged over designed positions that had been mutated by any design method. Plotted are mean and SEM values. Design protocols are colored as in panel D. C. Improvement in recapitulating evolutionary sequence profiles of RECON increases with the number of designed positions. For each benchmark set, the number of designed positions is plotted against the difference in evolutionary sequence similarity between RECON backbone minimized and MPI_MSD. Least-squares linear fit is shown, with an R-value of 0.61 and p value of 0.02. D. Difference in recapitulation of evolutionary sequence profile for the four largest benchmark sets by designs generated by RECON using fixed backbone (FBB) or backbone minimization (BBM) protocols, or MPI_MSD. P values were calculated using a paired two-tailed t test.