柠檬明串珠菌D-甘露醇合成途径关键酶的功能分析及代谢调控研究

As a naturally occurring functional sugar alcohol, mannitol is widely used in the food, pharmaceutical and chemical industries. The prevalent chemical synthesis process for commercial production of mannitol has a number of disadvantages including the low conversion rate and the byproduct formation. In order to improve the productivity of mannitol, many alternative processes based on the microbial fermentation have been suggested. . Leuconostoc citreum, hetero-fermentative lactic acid bacteria, is regarded as one of the most potential hosts for mannitol production. In the present study, we are to isolate the wild Leuconostoc citreum highly yeilding mannitol from natural fermented foods firstly, modify it using genetic engineering, improve the activity of key enzymes like mannitol dehydrogenase involved in mannitol synthesis, decrease the activity of fructokinase to block metabolic flux of the fructose to other directions, and thereby increase the conversion rate and productivity of mannitol. We also try to compare the wild and engineered strains, study the putative key enzymes involved in the process of fructose transport, fructose metabolism and mannitol conversion at the DNA level, and clarify interaction and regulation of these enzymes, metabolites and cofactors on the mannitol synthesis. The current study is to promote the research about the functional genomics of Leuconostoc citreum and modify the technique of improving Leuconostoc citreum function. This is an innovation and a driving force for the food fermentation industry and biotechnology.

甘露醇作为性能优异的功能性糖醇，已广泛应用于食品、医药、化工等行业。目前甘露醇生产常用的化学合成法，伴有转化率低、副产物产生等缺点。为提高甘露醇的产率，一直试图通过微生物发酵途径生产甘露醇。. 柠檬明串珠菌作为异型发酵乳酸菌的一种，被认为是最有潜力的甘露醇生产菌。本项目首先从自然发酵食品中筛选出高产甘露醇的野生柠檬明串珠菌，利用基因工程改造此菌株，提高甘露醇脱氢酶等甘露醇合成途径中关键酶活性，降低或失活果糖激酶等的活性，阻断果糖向其他代谢流的流向；构建菌株与野生型进行比较，针对其果糖吸收载体、代谢和甘露醇转化过程中的推测关键酶进行基因水平的功能研究，阐明这些酶、代谢产物和辅酶对甘露醇合成所起的调控作用。本项目研究将促进明串珠菌基因功能的研究，树立改造明串珠菌性能的方法和技术，这对食品工业与生物技术的发展是一个创新和推动。

甘露醇作为性能优异的功能性糖醇，已广泛应用于食品、医药和化工等行业。目前甘露醇生产常用的化学合成法，伴有转化率低、副产物产生等缺点。为提高甘露醇的产率，一直试图通过微生物发酵途径生产甘露醇。柠檬明串珠菌作为异型发酵乳酸菌的一种，被认为是最有潜力的甘露醇生产菌。因此，本项目首先从自然发酵食品中筛选出高产甘露醇的野生柠檬明串珠菌，对其甘露醇合成过程中关键酶的特性进行了研究，分析了关键酶、代谢产物对柠檬明串珠菌甘露醇细胞内外积累的调控作用，通过生物技术构建了高产甘露醇的柠檬明串珠菌，最后对柠檬明串珠菌产甘露醇的条件进行了优化，使柠檬明串珠菌甘露醇生产达到了最高。通过本项目的研究，有利于柠檬明串珠菌基因功能和表达调控研究的发展，有利于发展发酵法生产甘露醇的新技术，这将进一步降低甘露醇的生产成本并提高其市场竞争力，使甘露醇的应用前景更为广阔，同时提高柠檬明串珠菌在食品工业和生物技术中的利用价值，这对于食品发酵工业与生物技术的发展将是一个创新和推动。