链霉菌A01-chit33CT产纳他霉素和几丁质酶协同表达发酵条件优化  

Optimization of Fermentation Conditions of Collaborative Expression of Chitinase and Natamycin from Streptomyces lydicus A01-chit33 CT

作　　者：林振亚[1] 吴琼[1] 孙瑞艳[1] 陈捷[1] 李雅乾[1] 

机构地区：[1]上海交通大学农业与生物学院农业部都市农业(南方)重点实验室,上海200240

出　　处：《中国生物工程杂志》2012年第10期67-73,共7页China Biotechnology

基　　金：国家高技术研究发展专项经费(2011AA10A205);上海市科委重点项目(09391910900);农业部948项目(2011-G4);教育部高等学校博士学科点专项科研基金(20090073110048)资助项目

摘　　要：生物农药由于具有良好的生态效应和安全性,因此比化学农药更受到人们的青睐,生物农药的发展契合低碳、循环、清洁绿色经济发展理念。因此,寻求利于食品安全和环境保护,同时高效控制植物病害的新型生物农药成为时下及未来研究的热点。链霉菌以产生纳他霉素等抗生素起到生防作用。链霉菌株A01-chit33CT既可以产生纳他霉素又可以高表达几丁质酶活,生防效果大大增加。为确定链霉菌A01-chit33CT产纳他霉素和几丁质酶协同表达的发酵条件,初步探索了碳氮源和发酵条件对菌株产生纳他霉素和几丁质酶的影响。结果表明,葡萄糖促进纳他霉素的产生而抑制几丁质酶的表达,因此分两阶段添加葡萄糖和几丁质粉来达到二者协同表达。研究确定最佳发酵培养基为:葡萄糖40 g/L,几丁质粉10 g/L(发酵4 d添加),黄豆粉30 g/L,大豆蛋白胨10 g/L,CaCO35 g/L,MgSO4.7H2O 0.5 g/L,K2HPO40.5 g/L。最优发酵条件为:初始pH 6.0,温度28℃,转速180 r/min。在此条件下,链霉菌A01-chit33CT产纳他霉素达1.52 g/L,同时几丁质酶活达990 U/ml,二者比优化前的水平分别提高了1.95倍和2.27倍。Biological control of plant diseases is an effective method with the beneficial microorganisms or microbial metabolites against crop diseases.Streptomyces are already identified to produce a range of antibiotics including natamycin etc.,which exhibit an excellent activities against fungal pathogens.Streptomyces lydicus A01 has been proved as a biocontrol microbe with antagonistic activity against Botrytis cinerea by producing natamycin,which degrading the cell membrane of fungal pathogens.Natamycin,a polyene macrolide antibiotic with broad-spectrum antifungal activity,has a stable and strong inhibitory activity against many plant pathogenic fungi,and is commonly used as antifungal agent.Chitinase can degrade chitin and could be used for resist many fungal pathogens.In A01-chit33CT,natamycin was produced to inhibit plant pathogens via destructing fungal cell membrane,while chit33 plays an important role in hydrolyzing the chitin component in fungal pathogen cell wall.The coordination effect of natamycin and chit33 may be improved significantly. The optimal fermentation conditions for chitinase activity and natamycin produced in Streptomyces lydicus A01-chit33CT were studied.Firstly,the effect of different carbon and nitrogen sources on natamycin production and chitinase activities were investigated.The results showed that： glucose can promote natamycin production but inhibit the expression of chit33,so glucose and chitin powder were added into the fermentation medium with two-stage method to achieve collaborative expressions of both.The optimal medium compositions were as follow： chitin powder 10 g/L（after 4 days）,glucose 40 g/L,soybean meal 30 g/L,soya peptone 10 g/L,CaCO3 5g/L,MgSO4·7H2O 0.5g/L,K2HPO4 0.5 g/L.Secondly,the cultural conditions were further to optimum as follows： initial pH 6.0,temperature 28℃,rotation speed 180 r/min.The highest natamycin production could reach 1.52 g/L and chitinase activity reach 990U/ml under the optimal fermentation conditions.Both levels improved by 1.95 and 2.27

关 键 词：链霉菌A01-chit33CT 纳他霉素 几丁质酶 发酵 优化 

分 类 号：Q819[生物学—生物工程]