N-乙酰氨基葡萄糖(N-acetylglucosamine, GlcNAc)作为一种重要的功能性单糖,在缓解关节炎和治疗骨质疏松方面具有良好的生理功能。微生物发酵法是当前GlcNAc的主要生产方式,但常用的玉米浆干粉天然培养基成分复杂且稳定性差,会造成发酵批次之间产量的差异,不利于工业化生产。该研究以谷氨酸棒杆菌(Corynebacterium glutamicum)S9114为宿主,通过引入外源GlcNAc合成途径并阻断其分解途径,构建了GlcNAc生产菌株Cg5,在500 mL摇瓶中发酵72 h,GlcNAc产量达到14.4 g/L;其次,优化基因表达框架并采用启动子工程进一步优化关键酶的表达,获得了高效合成GlcNAc的重组菌株Cg10,其摇瓶发酵产量提高至28.8 g/L;之后,为了降低玉米浆干粉对发酵结果的影响,该研究分析了玉米浆干粉的成分,并通过响应面优化试验,确定了最适无机培养基组分。使用优化后的无机盐培养基,GlcNAc摇瓶产量达到27.5 g/L,与使用玉米浆干粉培养基的发酵结果相当。该研究通过构建高效合成GlcNAc工程菌株并采用无机培养基发酵的策略,为C.glutamicum在工业化生产GlcNAc中的应用提供了重要的工艺参数和理论支持。
N-acetylglucosamine (GlcNAc), an important functional monosaccharide, plays a significant role in alleviating arthritis and treating osteoporosis.Microbial fermentation is the primary method for GlcNAc production;however, the commonly used natural medium, corn steep liquor, has complex components and poor stability, which can lead to variations between fermentation batches, hindering industrial-scale production.In this study, Corynebacterium glutamicum S9114 was used as the host.By introducing an exogenous GlcNAc synthetic pathway and blocking its degradation pathway, a GlcNAc-producing strain named Cg5 was constructed.In a 500 mL shake flask fermentation for 72 h, the GlcNAc titer reached 14.4 g/L.Furthermore, by optimizing the gene expression framework and utilizing promoter engineering to enhance the expression of key enzymes, the recombinant strain Cg10 with high synthetic efficiency was constructed.The GlcNAc titer increased to 28.8 g/L in shake flask.Additionally, to minimize the impact of corn steep liquor on fermentation outcomes, the composition of corn steep liquor was analyzed, and the optimal inorganic medium components were determined through response surface methodology.Using the optimized inorganic medium, the GlcNAc titer reached 27.5 g/L, which was comparable to the results obtained with the corn steep liquor-based medium.Overall, this study provides important process parameters and theoretical support for the industrial application of C.glutamicum in GlcNAc production through the construction of high-efficiency recombinant strain for GlcNAc synthesis and the use of optimized inorganic medium for fermentation.
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