Fig. 3. Snapshots from EryAT6 simulations showing key residues with the active site.

a Wild-type EryAT6 shows the catalytic dyad His747 and Ser644, hydrophobic interactions between Tyr744, Val742, Leu673, and Val612, and transient charged interactions between Arg674, Asp612, and Asp743. Arg669 positions the carboxylate of the substrate for catalysis. The wild-type backbone is shown as a transparent overlay with all mutants. b The V742A mutation results in a weaker interaction between Ala742 and Tyr744 than in wild-type, as well as less bulkiness within the active site, but the overall structure remains similar to wild-type. c The Y744R mutant introduces transient charged interactions between Arg744, Asp743, and Asp613 while disrupting the wild-type hydrophobic interactions. d The double mutant V742A/Y744R combines the features of the two individual mutants, resulting in a much wider active site. e The L673H mutant results in π-stacking between Tyr744 and His673, creating a narrower active site and disrupting the interaction between Tyr744 and Val612. Time-averaged structures are shown Supplementary Fig. S5. Y744R mutation impacts the salt bridge, resulting in a larger inter-subunit distance. The wild-type model is overlaid as transparent ribbons. c Wild-type EryAT6 showing few interactions between Tyr744 and the large subunit. (d) Introduction of the L673H mutation results in a narrower active site via π-stacking between Tyr744 and His673. The wild-type model is overlaid as transparent ribbons.