Figure 3.

PseFDH CAST library design. (top) Active site of PseFDH variant A198G (black) highlighting residues selected for mutagenesis for switching cofactor specificity (green and violet), while recovering activity (blue). Residue R222 (black) was not mutated despite being suggested for mutagenesis by CRS-SALAD, as explained above. The carbon (yellow), nitrogen (blue), and oxygen (red) atoms of the cofactor are shown in stick format. The picture was generated with PyMol using PDB file 2NAD.²¹ (bottom) Amino acid (AA) alphabet, degenerate codon (N = A/T/G/C, K = T/G, D = A/G/T, B = C/G/T, W = A/T, R = A/G), and library size and coverage. The library size was calculated using the program CASTER 2.0.³¹