棘白菌素B高产菌株的构建及发酵调控

Construction and fermentation regulation of strains with high yields of echinocandin B

摘要:

棘白菌素B (echinocandin B,ECB)是第三代棘白菌素类抗真菌药物阿尼芬净的关键前体，是由构巢曲霉菌株发酵得到的次级代谢产物，其发酵效价受ECB合成通路及细胞形态的显著影响。本研究旨在通过对ECB生物合成过程中与转录激活、羟基化、细胞形态等相关的关键基因进行改造，以提高ECB的发酵效价，同时改变构巢曲霉细胞形态以降低发酵液黏稠度。结果表明过表达ecdB和ecdK基因使ECB发酵产量分别比野生菌株提高了25.8%和23.7%，达到(2 030.5±99.2) mg/L和(1 996.4±151.4) mg/L。但与细胞壁合成相关的fksA基因的缺失导致其细胞壁受损，影响了菌株生长和产物合成。对过表达ecdB的工程菌株进行50 L罐分批补料发酵调控，ECB发酵效价达到2 234.5 mg/L。本研究为后续菌株的代谢工程改造奠定了基础。

Abstract:

Echinocandin B (ECB) is a key precursor of the antifungal drug anidulafungin. It is a secondary metabolite of Aspergillus nidulans, and its titer in fermentation is significantly affected by the ECB synthesis pathway and cell morphology. In this study, the key genes related to the transcription activation, hydroxylation, and cell morphology during ECB biosynthesis were investigated to increase the fermentation titer of ECB and to change the cell morphology of Aspergillus nidulans to reduce the viscosity of the fermentation broth. The results indicated that after overexpression of ecdB and ecdK, the ECB titer increased by 25.8% and 23.7%, respectively, compared with that of the wild-type strain, reaching (2 030.5±99.2) mg/L and (1 996.4±151.4) mg/L. However, the deletion of fksA associated with cell wall synthesis resulted in damage to the cell wall, affecting strain growth and product synthesis. The engineered strain overexpressing ecdB was fermented in a 50-L bioreactor, in which the ECB titer reached 2 234.5 mg/L. The findings laid a research foundation for the subsequent metabolic engineering of this strain.