Deng Shiping, Lorito Matteo, Penttilä Merja, Harman Gary E
Department of Plant and Soil Sciences, Oklahoma State University, Stillwater, OK 74078, USA.
Appl Biochem Biotechnol. 2007 Jul;142(1):81-94. doi: 10.1007/s12010-007-0012-9.
Trichoderma is one of the most promising biocontrol agents against plant fungal diseases. In this study, a transgenic strain of Trichoderma atroviride was characterized. The transgenic strain contains an endochitinase gene (ThEn-42) driven by the cellulase promoter cbh1 of T. reesei for overexpression of ThEn-42. The culture filtrates of the transformant and the parental strain grown in eight different media were evaluated for chitinase and antifungal enzyme production based on activity gels, protein profiles, and antifungal activities. Results demonstrated that chitinases are important components and synergistic interactions play a key role in the antagonistic action of T. atroviride. Moreover, altering medium nutrient concentration and composition led to enhanced production of antifungal enzymes, a potential strategy for mass production. Two of the culture filtrates contained almost pure endochitinase, and could be excellent commercial sources for this enzyme. Several culture filtrates were highly antifungal. Two filtrates were so effective in biocontrol of a fungal pathogen, Penicillium digitatum, that they not only inhibited spore germination but destroyed the spores completely when 20 microl of culture filtrate (corresponding to approximately 104 microg of total protein) was applied in a total volume of 150 microl (approximately 0.7 mg protein ml(-1)).
木霉是最有前景的防治植物真菌病害的生物防治剂之一。在本研究中，对一株绿色木霉转基因菌株进行了特性分析。该转基因菌株含有一个由里氏木霉纤维素酶启动子cbh1驱动的内切几丁质酶基因（ThEn - 42），用于ThEn - 42的过表达。基于活性凝胶、蛋白质图谱和抗真菌活性，对在八种不同培养基中生长的转化体和亲本菌株的培养滤液进行了几丁质酶和抗真菌酶产生情况的评估。结果表明，几丁质酶是重要组成部分，协同相互作用在绿色木霉的拮抗作用中起关键作用。此外，改变培养基营养浓度和组成可提高抗真菌酶的产量，这是大规模生产的一种潜在策略。其中两种培养滤液几乎含有纯内切几丁质酶，可能是该酶的优良商业来源。几种培养滤液具有高度抗真菌性。两种滤液对指状青霉这种真菌病原体的生物防治效果非常好，当在150微升（约0.7毫克蛋白质/毫升）的总体积中加入20微升培养滤液（相当于约104微克总蛋白质）时，它们不仅抑制孢子萌发，还能完全破坏孢子。
