Extended Data Fig. 6. Principal component analyses identify amino acid chemical properties contributing to differential ligand perception.
a, Normalized ROS induction of flg22 variants (100 nM) in N. benthamiana fls2-1/2 expressing AtFLS2, SlFLS2, FcFLS2, VrFLS2. Each data point represents an average max relative light unit (RLU) from 4 plants with 4 leaf disks per plant. The values of each plate were normalized by adjusting maximum relative light unit (RLU) averages to a 0 (water) to 100000 (Pae) scale. One-way ANOVA and Dunnett’s multiple comparison were used to determine significance using the non-scaled data, p values are marked next to each bar. Error bars = SEM. At = Arabidopsis thaliana, Sl = Solanum lycopersicum, Fc = Fagus crenata, Vr = Vitis riparia. b, Principal component analysis (PCA) of FLS2 homologs and their associated residue properties. Scatterplot of contribution of chemical residue variables for top two PCA dimensions. Amino acid properties labeled in yellow display the highest variance and those in black display little variance between FLS2 homologs. c-d, Heatmap of bulkiness (top), hydrophobicity (middle), amino acid charge (bottom) along exposed residues of the LRR across FLS2 variants. All swapped residues in SynFcFLS213Qv (c) or SynVrFLS222XL (d) are labeled below the heatmaps. Swapped Residues with substantial change of chemical parameters are highlighted in bold (Δ bulkiness > ±2, Δ hydrophobicity > ±2, Δ charge > ± 0.5). The underlined residue is not directly located on the concave surface, yet was selected for engineering based on changes in side chain properties.