恩拉霉素生产菌株的遗传改造  

Genetic modification of an enramycin producing strain

作　　者：牟慧艳 刘扬[1] 王应东 刘宝爱 魏建军[2] 张会图[1] 

机构地区：[1]天津科技大学生物工程学院应用微生物与酶工程实验室,天津300457 [2]天津市新星兽药厂,天津300402

出　　处：《微生物学通报》2017年第1期126-132,共7页Microbiology China

基　　金：国家自然科学基金项目(No.81373309)~~

摘　　要：【目的】提高杀真菌素链霉菌发酵生产恩拉霉素的产量。【方法】利用定点突变技术,对恩拉霉素生产菌株杀真菌素链霉菌F1中影响细胞次级代谢及抗生素合成的核糖体S12蛋白的编码基因rps L进行改造,将第43位的赖氨酸(Lys)分别替换为天冬酰胺(Asn)和精氨酸(Arg),并对改造菌株L-M1(Asn43)和L-M2(Arg43)的生长特性、抗生素合成以及摇瓶发酵性能进行研究。【结果】与野生型菌株相比,改造菌株的生长特性及生理生化特性均发生了明显的改变:产孢周期明显缩短,野生型菌株在MS培养基中,28°C下需要培养5-7 d后才能产生孢子,而在相同条件下,改造菌株3 d后就能产生大量的孢子;恩拉霉素产量相对提高,摇瓶发酵条件下,改造菌株L-M1(Asn43)和L-M2(Arg43)的恩拉霉素产量分别可达到1 334 U/m L和1 456 U/m L,与野生型菌株F1相比分别提高了11.9%和22.1%。【结论】通过遗传改造,恩拉霉素的产量得到了提高,为其他位点的遗传改造提供了可行性。[Objective] To enhance the production of enramycin by Streptomyces fungicidicus. [Methods] In this study, the ribosomal S12 protein encoding gene rps L which affects the secondary metabolism and biosynthesis of antibiotics in enramycin producing strain of S. fungicidicus F1 was changed through the method of site-directed mutation, during which the Lys43 was substituted for Asn and Arg, respectively. And then the growth characteristics, synthesis of antibiotic and flask fermentation performance of the modified strains were studied. [Results] The results showed that the growth characteristics and physiological and biochemical characteristics of the modified strains were significantly changed compared with the wild type strain. Sporulation cycle was shorten, spore was produced from wild type strain using MS medium under the condition of 28 °C for 5-7 days, but the modified strains can produced large amounts of spore only after 3 days in the same conditions. Enramycin yield was relatively increased. Under the flask formation condition, the enramycin production of the modified strains of L-M1（Asn43） and L-M2（Arg43） can reach a maximum of 1 334 U/m L and 1 456 U/m L, respectively. Compared with the wild-type strain, it was improved by 11.9% and 22.1% separately. [Conclusion] Through the genetic modification, the yield of enramycin was improved, which might provide a feasibility for the modification of other sites.

关 键 词：恩拉霉素 抗生素 抗真菌素链霉菌 遗传改造 核糖体S12蛋白 定点突变 

分 类 号：Q93[生物学—微生物学]