高产α-酮戊二酸解脂亚洛酵母的选育及其发酵过程优化  

Breeding and process optimization of Yarrowia lipolytica for high-yieldα-ketoglutaric acid production

作　　者：房峻[1,2] 周景文 曾伟主[1,2] FANG Jun;ZHOU Jingwen;ZENG Weizhu(Key Laboratory of Industrial Biotechnology of Ministry of Education,School of Biotechnology,Jiangnan University,Wuxi 214122,China;Science Center for Future Foods,Jiangnan University,Wuxi 214122,China)

机构地区：[1]江南大学生物工程学院工业生物技术教育部重点实验室,江苏无锡214122 [2]江南大学未来食品科学中心,江苏无锡214122

出　　处：《食品与发酵工业》2021年第2期137-144,共8页Food and Fermentation Industries

基　　金：国家重点研发计划项目(2019YFA0904800);国家自然科学基金优秀青年基金项目(21822806)。

摘　　要：为了提高解脂亚洛酵母(Yarrowia lipolytica)发酵生产α-酮戊二酸(α-ketoglutaric acid,α-KG)的效率,对前期获得的1株α-酮戊二酸高产菌株1-C6进行了迭代诱变处理,并对最终获得的高产突变菌株进行了系统的发酵过程优化。首先,基于前期构建的高通量筛选方法,应用甲基磺酸乙酯(ethylmethylsulfone,EMS)、亚硝基胍(nitroso-guanidin,NTG)和紫外(ultraviolet,UV)等诱变方法对菌株1-C6进行迭代诱变,最终获得了1株高产突变菌株4-E2,其α-KG的产量相对出发菌株1-C6提高了56.7%。在3 L发酵罐水平上研究了搅拌转速、通风比、乙酸钠和CaCO 3添加量等条件对高产突变菌株4-E2积累α-KG的影响。结果表明,较高转速的两阶段通风比更适合于α-KG的生产,乙酸钠和CaCO 3的最佳添加量分别为6 g/L和10 g/L。在此基础上进行补料发酵策略的探索,确定了恒速连续补料发酵的策略,α-KG产量达75.86 g/L,相比分批发酵提高了44.4%。研究结果为促进发酵法生产α-KG的工业化提供了借鉴意义。In order to improve the fermentation intensity ofα-ketoglutaric acid(α-KG)with Yarrowia lipolytica,a highα-KG production strain 1-C6 screened previously was treated by iterative mutagenesis,and the fermentation conditions forα-KG production with the final obtained mutant were systematically optimized.Firstly,based on the previous established high-throughput screening method,a high-yield mutant 4-E2 was obtained by iterative mutagenesis with ethyl methane sulfonate,nitrosoguanidine and ultraviolet light.Compared with the starting strain 1-C6,the production ofα-KG was enhanced by 56.7%.Then,effects of agitation speed,ventilation ratio,addition of sodium acetate and CaCO 3 on the accumulation ofα-KG with mutant 4-E2 were investigated in 3 L fermenter.It was found that a two-stage ventilation ratio combined with relative higher rotating speed was more suitable forα-KG production,and the optimum addition of sodium acetate and CaCO 3 were 6 g/L and 10 g/L,respectively.On this basis,a strategy of constant feeding strategy was further determined after exploring different strategies of fed-batch fermentation.Under the optimal conditions,the production ofα-KG reached 75.86 g/L,which was 44.4%higher than that of the batch fermentation.The results could provide references for the industrialization ofα-KG fermentation.

关 键 词：解脂亚洛酵母 Α-酮戊二酸 诱变 高通量筛选 补料策略 

分 类 号：TQ921[轻工技术与工程—发酵工程]