Fig. 2: DMS screen reveals D136 and D139 as key residues on FDX1 regulating cuproptosis.

A Heatmap of the results of the DMS screen in ABC1 cells. Colors are plotted as a function of the mean log₂ fold change (LFC) in viability of duplicates scaled relative to the mean viability scores obtained from the nonsense (amino acid X) and silent mutation distributions (Fig. 1D). The redder the color, the higher the viability. The gray bars indicate either no mutation (i.e., original amino acid) or missing data acquisition. The secondary structures of specific segments of FDX1 are indicated; the third alpha helix is emphasized. Residues within clusters of mutational intolerance are emphasized. Amino acids are colored coded by their chemical properties. The average ΔΔG, which predicts a mutation’s impact on protein stability, is indicated at each position on FDX1. B The left scatterplot shows the average viability LFC and average positional ΔΔG for each FDX1 amino acid position when mutated to all possible residues. The right panel shows the same, but only when mutated to positive/basic residues (arginine, lysine, histidine). Indicated in orange are the Fe-S cluster binding cysteines; and in magenta, residues D136 and D139. C Crystal structure of human FDX1 PDB: 3P1M in complex with an Fe-S cluster shown in spheres. Residues D136 and D139 are presented in magenta sticks and the four cysteines that coordinate the Fe-S cluster in yellow sticks. Transparent surface model is shown to highlight solvent-exposure of D136 and D139.