基因工程菌EGs2产β-葡聚糖酶条件优化及产酶特性  

Optimization of Recombinant Escherichia coli EGs2 Fermentation Condition Affecting β-glucanase Production and Its Characteristics of Enzyme

作　　者：李青青[1] 陈启和[1] 蒋孝燕[1] 张秀艳[2] 李婷[1] 何国庆[1] 

机构地区：[1]浙江大学食品科学与营养系,杭州310029 [2]华中农业大学食品安全与微生物学系,武汉430070

出　　处：《农业生物技术学报》2007年第4期708-712,共5页Journal of Agricultural Biotechnology

基　　金：国家自然科学基金(No.20276064)资助

摘　　要：用乳清粉作为主要培养基组成,采用正交试验对大肠杆菌(Escherichia coli)重组菌EGs2的发酵条件进行了优化,并对粗酶液的酶学性质进行了研究。重组菌EGs2的最佳发酵条件为:床转速170r/min,接种量1%,发酵培养基初始pH 6.0,种龄12h。重组菌EGs2 β-葡聚糖酶的表达与菌体生长呈正相关,发酵14h后,菌体生物量最高可达1.71g/L,β-葡聚糖酶活力最高可达321.56U/mL。所得粗酶液的最适反应温度60℃,pH 6.0,在70℃以下保温20min后,残余酶活力均在80%以上。在pH 5.0～9.0放置48h后仍能保持均90%以上的残余酶活力。The fermentation conditions of mutant EGs2 obtained from directed evolution for thermostable β-glucanase for Escherichia coli β-glucanase production were investigated by orthogonal experiment. Fermentation was conducted in 250 mL flask, each containing 30 mL of medium contained whey 10 g/L, yeast extract 5 g/L, NaCl 10 g/L, the temperature was 37℃. The optima culture conditions were as following： initial pH 6.0, shaking speed 170 r/min, inoculation volume 1% and inoculation time 12 h. β-glucanase production by mutant EGs2 was associated with cell growth and biomass. β-glucanase activity was increased significantly when cells entered growth phase. The bacterium began the stable phase after cultured for 14 h with the highest biomass 1.71 g/L and β-glucanase activity 321.56 U/mL. When β-glucanase was used as a substrate, the optimum temperature and pH were 60 ℃ and 6.0, respectively. After 20 min incubation at 40, 50, 55, 60, 65 and 70 ℃ respectively, the residual activity remained at least 80%. After 48 h conservation at pH 5.0-9.0, the residual activity remained at least 90%. The enzyme was stable below 70 ℃ and at pH 5.0-9.0.

关 键 词：Β-葡聚糖酶 重组菌EGs2 发酵条件 酶学性质 

分 类 号：S182[农业科学—农业基础科学]