链霉素抗性选育ε-聚赖氨酸高产菌株

Breeding of High ε-poly-L-lysine-producing Strain by Streptomycin Resistance

摘要:

基于核糖体工程的育种方法，通过逐级赋予小白链霉菌M-Z18链霉素抗性，以提高产生菌的ε-聚赖氨酸（ε-PL）合成能力。首先，通过筛选低浓度链霉素（1~5 MIC）抗性突变株，将M-Z18的ε-PL的摇瓶产量从1.6提高到2.43 g/L；随后，再次提升菌株的链霉素耐受性（5~10 MIC），进一步增强菌株的ε-PL合成能力；最后，获得一株高产突变菌SS-19，其ε-PL产量和单位菌体合成能力分别为3.13 g/L和0.58 g/g，较出发菌M-Z18分别提高了95.63%和137.61%。研究表明，SS-19的中心碳代谢途径及ε-PL合成相关的关键酶活性有所增加，意味着ε-PL的合成代谢被明显加强。在以葡萄糖为碳源的RSM培养基中，SS-19比前期育种获得的高产菌株表现出更好的ε-PL合成能力。以上结果表明，通过逐步引入高浓度链霉素抗性的筛选方法可有效提升小白链霉菌的ε-PL产量。

Abstract:

Using a ribosome engineering-based breeding approach, the ε-Poly-L-lysine (ε-PL) biosynthesis capability of Streptomyces albulus M-Z18 was gradually enhanced by increasing streptomycin resistance in a stepwise manner. Firstly, the ε-PL production by S. albulus M-Z18 was increased from 1.6 to 2.43 g/L through screening low streptomycin-resistant (1~5 MIC) mutant strains. Subsequently, the streptomycin tolerance was further increased (5~10 MIC), to enhance ε-PL biosynthesis capability. Finally, a high ε-PL-producing mutant S. albulus SS-19 was obtained, with the yield of ε-PL and the unit cell synthesis capability as 3.13 g/L and 0.58 g/g, respectively, which were increased by 95.63% and 137.61%, respectively, compared with the original strain M-Z18. Results showed that the activities of the key enzymes involved in the central carbon metabolic pathway and ε-PL biosynthesis pathway were significantly enhanced, suggested that the anabolic metabolism of ε-PL was significantly enhanced. In the RSM medium with glucose as the carbon source, S. albulus SS-19 exhibited a greater ε-PL biosynthesis capability compared with the high-yield strains obtained from previous breeding. The above results indicated that the ε-PL production by Streptomyces albulus could be improved effectively by the screening method involving the gradual increase of streptomycin resistance.