Serratia sp. TKU016所生產蛋白酶及幾丁聚醣酶之純化與定性

Purification and Characterization of Protease and chitosanase from Serratia sp. TKU016

張韜仁(Tao-Jen Chang)

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.2008.00856

摘要

TKU016 係以蝦殼粉為唯一碳/氮源，篩選自台灣北部土壤之一株幾丁聚醣酶及蛋白酶生產菌，經鑑定為Serratia sp.。TKU016生產幾丁聚醣酶及蛋白酶之較適培養條件為含有1% 蝦殼粉、0.1 % K2HPO4及0.05 %MgSO4．7H2O之液態培養基(pH 7)在30℃搖瓶培養3天。所得發酵上清液，經硫酸銨沉澱、DEAE-Sepharose及Sephacryl S-100等層析步驟，可分離出一種幾丁聚醣酶及一種蛋白酶，經SDS-PAGE測其分子量分別為 65 kDa及53 kDa。於最適反應pH、最適反應溫度、pH安定、熱安定性方面，幾丁質酶的條件為pH 7、50℃、pH 4-8及<80℃；蛋白酶的為pH 11、40℃、pH 5-11及<90℃。幾丁聚醣酶活性受Mn2+所抑制。蛋白酶對偶氮白蛋白、偶氮酪蛋白為基質時，具有較佳的活性，對於凝膠蛋白、纖維蛋白、彈性蛋白的活性不佳。

關鍵字

幾丁質聚醣酶 ； 幾丁質酶 ； 蛋白酶

並列摘要

The chitosanase and protease producing strain, Serratia sp. TKU016, was isolated from the soil in Taiwan. The optimized condition for chitosanase and protease production were found when the culture was shaken at 30℃for 3 days in 100mL of medium contain 1% shrimp shell powder (SSP), 0.1 % K2HPO4 and 0.05 % MgSO4．7H2O (pH8). One chitosanase and one protease were purified by chromatography procedures of DEAE-Sepharose, and Sephacryl S-100. The molecular mass of the chitosanase and protease determined by SDS-PAGE was approximately 65 kDa and 53 kDa, respectively. The optimum pH, optimum temperature, pH stability, and thermal stability of chitosanase were pH 7、40℃、pH 4-8 and <80℃, respectively. The optimum pH, optimum temperature, pH stability, and thermal stability of protease were pH 11、40℃、pH 5-11 and <90℃,respectively. The chitosanase was inactivated by Mn2+. The protease showed good activity toward azocasein and azoalbumin as substrates, low activity with gelatin, fibrin, and elastin.

並列關鍵字

Serratia sp. ； chitosanase ； chitinase ； protease

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