Production and characterization of chitinolytic enzymes from microorganisms

Mucor sp. (TISTR 3301) and Paecilomyces lilacinus (KD-1-1) were found to effectively produce chitinolytic enzymes in medium P1-5. In aerobic culture, 4.10-4.42 U/ml of chitinase and 4.27-5.73 U/ml of chitobiase were produced by Mucor sp. (TISTR 3301) on day 4 and 1.88-2.16 U/ml of chitinase, 51.00-54.20 U/ml of chitobiase were produced by P. lilacinus (KD-1-1) on day 2. Actinomycete (1-3) was grown in medium P2, 1.76-2.60 U/ml of chitinase and 8.40-9.20 U/ml of chitobiase were respectively obtained. After mutagenesis of P. lilacinus (KD-1-1) using NTG, the ability of the mutants MKD-04, M3-17 and M3-44 to produce chitinolytic enzymes was improved and 8.36-13.30 U/ml of chitinase and 195.50-427.50 U/ml of chitobiase were produced by the mutants. Optimum conditions for the activities of the enzymes produced by Mucor sp. (TISTR 3301) were the pH of 5.0 for chitinase and chitobiase and the temperatures of 50 deg C and 60 deg C respectively. Both enzymes were found stable at pH 4.0-6.0 and at the temperature below 30 deg C. For P. lilacinus (KD-1-1), chitinase and chitobiase were most active at pH 5.0, the optimum temperatures of chitinase and chitobiase were 40 deg C and 50 deg C, respectively. Both enzymes were found stable at a pH range of 5.0-6.0 and at temperature below 40 deg C. The optimum pH and temperature of chitinase and chitobiase from Actinomycete (1-3) were pH 6.0, 5.0 and 40 deg C, 50 deg C, respectively. Both enzymes were found stable at a pH range of 5.0-7.0 and at temperature below 30 deg C. Swollen chitin degradation by chitinolytic enzymes from both fungi produced chitobiase and NAG. Except for Actinomycete (1-3), only NAG was separated from HPLC analysis. Partial purification of chitinolytic enzymes was carried out by (NH4)2SO4 precipitation followed by gel filtration. After gel filtration, chitinases of Mucor sp. (TISTR 3301), P. lilacinus (KD-1-1) and Actinomycete (1-3) were purified to 5.76, 4.66 and 9.9 times respectively. The chitobiases were purified to 8.99, 2.48 and 6.14 times for Mucor sp. (TISTR 3301), P. lilacinus (KD-1-1) and Actinomycete (1-3) respectively.