FIGURE 5.

Degradation of chitosan with wild-type and mutant ChiAs. A10 mg/ml chitosan solution in 40 μm sodium acetate, 100 μm NaCl, pH 5.5, and 0.1 mg/ml bovine serum albumin was incubated with enzyme (5 μg of enzyme/10 mg of chitosan). Degradation was allowed to proceed for increasing time intervals. The reactions were stopped by lowering the pH to 2.5 by the addition of 1.0 m HCl and immersing the samples in boiling water for 2 min. The fraction of cleaved glycosidic bonds (α) at different time points during the reaction was determined by NMR, as described under “Experimental Procedures.”