Fig. 3: The side chains of Tyr110 and Lys105 do not affect the pK_a-value of the ScGrx7 active site thiol group.

Freshly reduced wild-type ScGrx7 (black symbols) and selected mutants (colored symbols) were incubated with 150 µM iodoacetamide at 23 °C for 180 s in a three-buffer system at pH 3.5–8.5. Residual activities were measured in a standard HEDS assay and normalized against mock controls that were incubated in parallel without iodoacetamide. a Data for wild-type ScGrx7 (WT) and the mutants E170A, Y110A, K105A, and K105E. Samples containing iodoacetamide were apparently more stable at lower pH values than the mock controls resulting in residual activities slightly above 100%. The pK_a values from the sigmoidal fits are indicated. Data points and error bars represent the mean ± s.d. of three independent experiments. b Summary and statistical analyses of the pK_a values from panel (a). Sigmoidal fits using the four parameter Hill function and P values from one way ANOVA analyses followed by a Holm–Sidak test were calculated in SigmaPlot 13 (P > 0.05: ns; P ≤ 0.001: ***). Source data are provided as a Source Data file.