FIGURE 4.

Activity and binding measured with α-chitin. Wild type ChiA (♢), ChiA-W275A (▵), and ChiA-W167A (□) are shown. The concentration of α-chitin was 4 mg ml⁻¹ except in the binding experiments with ChiA-W275A and ChiA-W167A, where it was 2 mg ml⁻¹. The concentration of ChiA was 20 nm. Reactions were supplemented with chitobiase (20 nm), and the release of reducing groups (expressed in NAG₂ equivalents) in time was followed. A, hydrolytic activity in the short to medium time scale experiments. B, concentration of free enzyme in the short to medium time scale experiment. C, hydrolytic activity in the long time scale experiments.