L-丝氨酸在医药、食品和化工等领域应用广泛,然而L-丝氨酸是目前工业化生产难度较大的氨基酸之一。该文以实验室前期构建的产L-丝氨酸的谷氨酸棒杆菌A36-AB为出发菌株,利用常温常压等离子体诱变(atmospheric and room temperature plasma,ARTP)结合高通量筛选,获得突变菌株A36-AB-pDser-6。突变菌株前期生长迅速,60 h时L-丝氨酸产量为15.60 g/L。进一步利用响应面法优化突变菌株发酵培养基,L-丝氨酸产量达到29.23 g/L,生产强度为0.48 g/(L·h),生产强度较出发菌株[0.36 g/(L·h)]提高了33%;发酵周期从120 h缩短至60 h。对出发菌株及高产突变株进行全基因组测序及比较分析,发现突变株存在12个单碱基突变(single nucleotide variant,SNV)和15个插入缺失突变(insertions and deletion,InDel)。其中糖酵解途径中的关键酶葡萄糖-6-磷酸异构酶(由pgi基因编码)发生较多突变,这可能是由于突变菌株发酵周期缩短。该研究为利用谷氨酸棒杆菌产L-丝氨酸的工业化生产奠定了基础。
L-serine is widely used in industries such as medicine, food, and chemicals, but it remains one of the amino acids with a high production challenge for industrial-scale synthesis.This study used Corynebacterium glutamicum strain A36-AB, which was previously engineered in our lab for L-serine production, as the starting strain.Atmospheric and room temperature plasma (ARTP) mutagenesis, coupled with high-throughput screening, was employed to generate the mutant strain A36-AB-pDser-6.The mutant strain exhibited rapid growth, achieving an L-serine yield of 15.60 g/L after 60 hours of fermentation.Further optimization of the fermentation medium for the mutant strain was conducted using response surface methodology, leading to an enhanced L-serine yield of 29.23 g/L and a productivity of 0.48 g/(L·h), a 33% increase over the starting strain’s productivity of 0.36 g/(L·h).Moreover, the fermentation cycle was reduced from 120 hours to 60 hours.Whole-genome sequencing and comparative genomic analysis of the starting and high-yield mutant strains revealed 12 single nucleotide variants (SNV) and 15 insertions and deletions (InDel) in the mutant strain.Notably, mutations were observed in the key glycolytic enzyme phosphoglucose isomerase (encoded by pgi gene), which may account for the shortened fermentation cycle.This work provides a foundation for the industrial-scale production of L-serine using Corynebacterium glutamicum.
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