加强表达碳代谢全局转录因子对谷氨酸棒杆菌产l-丝氨酸的影响

Effects of overexpression of global transcription factors of carbon metabolism on l-serine production by Corynebacterium glutamicum

摘要:

【背景】谷氨酸棒杆菌(Corynebacterium glutamicum)广泛应用于氨基酸的生产，是l-丝氨酸的主要生产菌株之一。全局转录因子在胞内代谢物合成的调节中发挥着重要的作用，调节具有特定功能的转录因子可以有效增加目标氨基酸的合成。【目的】研究谷氨酸棒杆菌碳代谢全局转录因子GlxR、RamA、RamB、LldR、GntR1、SigB和SigH对谷氨酸棒杆菌A36-AC的生长和l-丝氨酸产量的影响。【方法】以本实验室保藏的产l-丝氨酸的谷氨酸棒杆菌A36-AC为出发菌株，构建单独加强表达转录因子基因glxR、ramA、ramB、lldR、gntR1、sigB和sigH的重组菌，通过发酵考察重组菌的生长和产l-丝氨酸情况，并对结果进行分析。【结果】在谷氨酸棒杆菌A36-AC中加强表达RamB和GlxR可以促进l-丝氨酸的合成且不影响菌株A36-AC的生长，而加强表达转录因子RamA、LldR、GntR1、SigB和SigH对菌株A36-AC的生长和l-丝氨酸产量均无明显影响。进一步将2种转录因子RamB和GlxR进行组合加强表达，构建重组菌株AC-pZM1-ramB-glxR，发酵结果表明，重组菌最大OD₅₆₂为30.3，而出发菌株A36-AC最大OD₅₆₂为34.1；重组菌株的l-丝氨酸产量和生产强度分别达到43.10 g/L和0.36 g/(L·h)，比出发菌株A36-AC分别提高15.9%和16.1%。【结论】在谷氨酸棒杆菌中加强表达转录因子RamB和GlxR可以促进l-丝氨酸的合成，本研究为进一步提高微生物发酵生产l-丝氨酸的水平奠定了基础。

Abstract:

[Background] Corynebacterium glutamicum is widely used for amino acid production and it is one of the major l-serine-producing strains. Global transcription factors play an essential role in the regulation of intracellular metabolite synthesis, and the regulation of transcription factors with specific functions can effectively increase the synthesis of target amino acids. [Objective] To systematically investigate the effects of overexpressing the global transcription factors (GlxR, RamA, RamB, LldR, GntR1, SigB, and SigH) of C. glutamicum carbon metabolismon on the growth of C. glutamicum A36-AC and l-serine titer.[Methods] With C. glutamicum A36-AC capable of producing l-serine as the starting strain, recombinant strains with individual overexpression of the transcription factor genes glxR, ramA, ramB, lldR, gntR1, sigB, and sigH were constructed. The growth and l-serine titer of the recombinant strains were examined by fermentation and the experimental results were analyzed. [Results] The overexpression of RamB and GlxR in C. glutamicum A36-AC promoted the synthesis of l-serine and did not affect the growth of C. glutamicum A36-AC, whereas the overexpression of RamA, LldR, GntR1, SigB, and SigH had no significant effects on the growth of C. glutamicum A36-AC or the l-serine titer. RamB and GlxR were overexpressed in A36-AC, and the recombinant strain AC-pZM1-ramB-glxR was obtained. The fermentation profiles indicated that the maximum OD₅₆₂ of AC-pZM1-ramB-glxR was 30.3, while that of C. glutamicum A36-AC was 34.1. The l-serine titer and productivity of the recombinant strain AC-pZM1-ramB-glxR reached 43.10 g/L and 0.36 g/(L·h), respectively, which were 15.9% and 16.1% higher than those of C. glutamicum A36-AC. [Conclusion] The overexpression of RamB and GlxR in C. glutamicum promotes the synthesis of l-serine. The findings lay a foundation for further improving the production of l-serine by microbial fermentation.