- 期刊
篩選幾丁質分解酶生產菌與其酵素之特性分析
Screening of a Chitinase-Producing Strain and Characterization of Its Chitinases
摘要
本研究自台北虎山溪之土壤中，篩選出具有生產幾丁質分解酶(chitinase)之菌株，此菌株為革蘭氏陰性桿菌，經食品工業研究所鑑定為Aeromonas hydrophila，命名為Aeromonas hydrophila DYU-Too10。此菌株於CB(chitin broth)培養基中，振盪培養120 h，幾丁質分解酶活性可達316U/L，其水解產物主要以N-乙醯葡萄糖胺(N-acetylglucosamine)為主。分析粗酵素特性，發現此酵素於酸性環境中易喪失活性，對熱敏感，其最適反應溫度為40℃，最適反應之pH為7.0。以不同碳源培養此菌株，得知以2%(w/v)粉狀幾丁質作為碳源之酵素產量為最高，而利用其他醣類作為碳源時，菌株則無法大量分泌幾丁質分解酶，推測其所分泌之幾丁質分解酶是由幾丁質誘導產生的。將粗酵素液經硫酸銨沉澱、透析、DEAE-Sepharose CL-6B及Sephadex G-100純化後，可獲得具有幾丁質分解酶活性之蛋白質。經活性電泳染色法證實，有三種幾丁質分解酶存在，其分子量分別為40、50及55 kDa。
並列摘要
In this study, a chitinase-producing strain was isolated from soil in the Hu-Shan Creek in Taipei County, Taiwan. The Food Industry Research and Development Institute (Hsinchu, Taiwan) identified this strain as Aeromonas hydrophila, a gram-negative, rod-shaped bacterium (designated as Aeromonas hydrophila DYU-Too10). The activity of the chitinases produced by the strain was 316 U/L after 120 h when cultured in Medium CB (chitin broth). The main hydrolysate of chitin obtained with the aid of chitinases was N-acetylglucosamine. The crude enzyme reacted optimally at 40℃, being very sensitive to thermal heat. Having an optimal pH of 7.0, this enzyme's activity decreased rapidly in an acidic environment. Various carbon sources were used to cultivate this strain in order to study the effect of a carbon source on the chitinase activity. The experimental results indicated that the chitinases produced by this strain were mainly induced by chitin and its derivatives. A protein having chitinase activity was purified by a series of procedures including ammonium sulfate precipitation, dialysis, anion exchange of DEAE-sepharose CL-6B and filtration of sephadex G-100. Three chitinases having molecular weights of 40, 50 and 55 kDa, respectively, were identified by a gel activity-staining method.