Figure 1.

pH, temperature optima, kinetic parameters and activity on different insoluble substrates of EcChi1. The pH and temperature optima were determined by incubating 30 mg/ml colloidal chitin and 30 μg of EcChi1 for 1 h at 40 °C in various buffers at different pH values, as follows: (A) 50 mM glycine-HCl (pH 2.0, ▼), 50 mM sodium citrate (pH 3.0–6.0, ■), 50 mM sodium acetate (pH 4.0–6.0, ▲), 50 mM sodium phosphate (pH 6.0–8.00, ●), 50 mM Tris-HCl (pH 8.0–9.0, ♦) and 50 mM glycine-NaOH (pH 9.0–10.0, ○) and at 10 °C to 80 °C (B), Different concentrations of colloidal chitin (2.5–60 mg/ml) and 30 μg of EcChi1 in 50 mM sodium citrate buffer, pH 5.0. The average of the triplicate data was fitted to the Michaelis-Menten equation by a nonlinear regression function of graphpad prism version 6.0 (C), The reaction mixture containing 30 μg EcChi1 and 1 mg ml⁻¹ of one of the polymeric substrates, was incubated at pH 5.0 and 40 °C for 1 h. The reaction mixture was centrifuged, and products were quantified using a reducing group assay (D). The error bars represent the standard deviations from three individual experiments.