Effects of π-helix mutations on PccGCS characteristics. (A) Enzyme kinetics of PccGCS WT (WT) and H237A/K238A (HK). Red values represent proteins in the ligated Fe(II)-O₂ state, and black values represent proteins in the unligated Fe(II) state. Turnover numbers were calculated from assays at V_max conditions with varying enzyme concentrations, and error bars represent standard deviations. (B) Analytical gel filtration analysis of oligomerization states of PccGCS WT (solid line) and H237A/K238A (dotted line). Different oligomeric states are annotated for both constructs on the graph. The molecular weights and retention times of globular protein standards are plotted above the graph. (C) Circular dichroism spectra for PccGCS WT (solid line) and H237A/K238A (dotted line). The plots are the averages of three scans for each construct.