一株高产脯氨酸的嗜醋酸棒杆菌的选育及发酵条件优化

Screening and Fermentation Optimization of Corynebacterium acetoacidophilum Mutant for High-level Production of L-proline

以嗜醋酸棒杆菌为出发菌株,经过片段化全基因组体外诱变、重组和连续的磺胺胍抗性筛选,获得一株L-脯氨酸的高产菌株。摇瓶发酵优化结果表明,葡萄糖、生物素和硫胺素的最适用量分别为16%、300μg/L、400μg/L,最适pH为6.8～7.0,装液量为25ml/500ml摇瓶,发酵培养72h后L-脯氨酸产率高达到75.6g/L,与对照相比提高了5%。考察了50L发酵罐中细胞生长对L-脯氨酸产量的影响,补料分批发酵结果表明(比生长速率分别为0.06/h、0.08/h和0.1/h),比生长速率在0.08/h左右时L-脯氨酸的产率最高,L-脯氨酸的比生产速率QP达到0.091g/(g.h),产率高达82.1g/L,比优化前提高了14%。

Corynebacterium acetoacidophilum was used as the original strain for stepwise mutation by whole genome fragmentation mutation（UV and NaNO2）and subsequent sulfaguanidine resistance screening.A L-proline over-producing mutant was obtained.The optimum contents of glucose,biotin and thiamin in shake-flask fermentation were 16%,300μg/L and 400μg/L respectively,and pH is 6.8~7.0,where 25ml medium was cultured in 500ml shake-flask.After 72h by flask-shaking batch fermentation,the production of L-proline reached 75.6g/L,5% higher than with the control.To investigate the effect of cell growth on the L-proline production,50L fed-batch culture was performed at the specific growth rate（μ）of 0.06/h,0.08/h and 0.1/h.The results showed that the specific L-proline production rate reached 0.091g/（g.h）and the highest production was 82.1g/L,increased by 14% compared with the control when the specific growth rate（μ）was 0.08/h.