Bacillus cereus TKU022所生產幾丁聚醣酶及蛋白酶之特性分析與應用

Abstract

The chitosanase -producing bacterium, Bacillus cereus TKU022, was isolated from soil in the north Taiwan by using shrimp head powder as the sole carbon/nitrogen source. The supernatant of the culture medium contains the chitosanase and the protease the activity. The optimized condition for chitosanase production was found when the culture was shaken at 37°C for two days in 50mL of medium containing 1.5% SHP, 0.1% K2HPO4, 0.05 % MgSO4．7H2O ( pH 5). The chitosanase and protease were purified from the culture supernatant by ammonium sulfate precipitation, DEAE-Sepharose, Phenyl Sepharose, Macro-Prep DEAE and Sephacryl S-100. The molecular masses of the chitosanase and protease determined by SDS-PAGE were approximately 44 and 45 kDa, respectively. The optimum pH, optimum temperature, pH stability and thermal stability of TKU022 chitosanase and protease were pH 7, 60 ℃, pH 7-10, <40 ℃and pH 10, 50-60 ℃, pH 6-10, <50 ℃, respectively. The chitosanase activity was inhibited by Cu2+ and Mn2+, but not by Tween 20, Triton X-100 (nonionic surfactant) and SDS (anionic surfactant).The protease activity was inhibited by Mn2+, EDTA and SDS, but retained 97 %, 105 % and 94% of its original activity in the presence of 2 % Triton X-100, 2 % Tween 20 and 2 % Tween 40, respectively. B. cereus TKU022 was incubated for 8 days under the optimized culture conditions（1.5 % SHP, 37 ℃, pH 5）and analyzed the reducing sugar. TKU022 culture supernatant incubated for 5 days had the highest reducing sugar（649 mg/mL）and enhanced the growth rate of Lactobacillu paracasei TKU010 in MRS broth.