Serratia sp. TKU017所生產蛋白酶及幾丁質酶之純化與定性

Purification and Characterization of Protease and Chitinase from Serratia sp. TKU017

李正宇(JENG-YU LI)

指導教授 ： 王三郎

淡江大學/理學院/生命科學研究所碩士班/碩士(2008年)

https://doi.org/10.6846/TKU.2008.00864

摘要

TKU017 係以蝦殼粉為唯一碳/氮源，篩選自台灣北部土壤之一株蛋白酶及幾丁質酶生產菌，經鑑定為 Serratia sp.。TKU017 生產蛋白酶及幾丁質酶之較適培養條件為含有 2% 蝦殼粉、0.1% K2HPO4 及 0.05% MgSO4．7H2O 之 100 mL 液態培養基 (pH 5) 在 25℃ 搖瓶培養 3 天。所得發酵上清液，經硫酸銨沉澱、DEAE-Sepharose、Phenyl-Sepharose及Sephacryl S-100等層析步驟，純化出一種蛋白酶及一種幾丁質酶，經 SDS-PAGE 測其分子量分別為 53 kDa 及 65 kDa。於最適反應 pH、最適反應溫度、pH安定、熱安定性方面，蛋白酶的為 pH 9、40℃、pH 5-11 及 ＜40℃；幾丁質酶的為 pH 5、50℃、pH 5-7 及 ＜50℃。蛋白酶活性受 EDTA 所抑制，為金屬型蛋白酶，幾丁質酶活性則受 Cu2+、Mn2+、SDS 所抑制。

關鍵字

Serratia sp. ； 蛋白酶 ； 幾丁質酶 ； 蝦殼粉

並列摘要

The protease and chitinase producing strain, Serratia sp. TKU017, cultured by using shrimp shell powder as the only carbon/nitrogen source, was isolated from the soil in the northern Taiwan. The optimized condition for the production of the protease and chitinase was found when the culture was shaken at 150 rpm at 25℃for 3 days in 100mL of medium contain 2% shrimp shell powder (SSP), 0.1 % K2HPO4 and 0.05 % MgSO4．7H2O (pH5). One protease and one chitinase were purified by chromatography procedures of DEAE-Sepharose, Phenyl-Sepharose, and Sephacryl S-100. The molecular mass of the protease and the chitinase determined by SDS-PAGE was approximately 53 kDa and 65 kDa, respectively. The optimum pH, optimum temperature, pH stability, and thermal stability of protease were pH 9, 40℃, pH 5-11, and ＜40℃, respectively. The optimum pH, optimum temperature, pH stability, and thermal stability of chitinase were pH 5, 50℃, pH 5-7, and及＜50℃, respectively. The protease activity was inactivated by EDTA；The chitinase activity was inactivated by Cu2+, Mn2+ and SDS.

並列關鍵字

Serratia sp. ； protease ； chitinase ； shrimp shell powder

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