嗜热厌氧产乙醇杆菌乙醇代谢途径的初步研究

以乙醇产量和细胞密度的提高为目标,从碳源和氮源入手,对嗜热厌氧产乙醇杆菌THermoanaerobacter ethanolicus JW200的培养基作了初步优化以提高乙醇代谢途径中的关键酶的得率,便于提纯.葡萄糖浓度的提高对乙醇的产生有明显的诱导作用,但当葡萄糖浓度高于2%,乙醇产量反而下降.酵母粉对乙醇产量没有促进作用,单纯提高酵母粉浓度对细胞密度无显著影响,而同时提高葡萄糖和酵母粉浓度至2.0%,OD600最高可达2.0.2%葡萄糖,0.5%酵母粉对单位细胞乙醇产量的诱导效果最佳.T.ethanolicusJW200粗酶液中未检测到丙酮酸脱羧酶的活性,经亲和柱Cibacron Blue-3GA部分纯化,在NAD洗脱蛋白中检测到辅酶A依赖型乙醛脱氢酶的活性,表明辅酶A依赖型乙醛脱氢酶是该菌乙醇代谢途径中的关键酶,它和乙醇脱氢酶共同作用,将乙酰辅酶A转化成乙醇.

Primary Study on Ethanol Production Pathway in Thermoanaerobacter ethanolicus

In order to induce the key enzymes of ethanol production pathway in ethanolicus JW200 for purification, the paper optimized the medium to increase the ethanol and cell yields. Final ethanol production was obviously enhanced when increasing concentration of glucose, but no progress was detected when glucose reached higher than 2%. No greater ethanol production or cell biomass was observed if simply increase concentration of yeast extract. The maximum OD6oo reached 2.0 when increasing glucose and yeast extract to 2.0%. The best ethanol induction medium contained 2.0% glucose and 0. 5% yeast extract. Cell-free extracts were prepared from T, ethanolicus JW200, and no pyruvate decarboxylase activity was detected in it. After partially purified by affinity chromatography Cibacron Blue- 3GA, CoA-acylating aldehyde dehydrogenase activity was detected in NAD elution proteins. The results indicated CoA-acylating aldehyde dehydrogenase was the key enzyme in ethanol production pathway of T. ethanolicus JW200, which convert acetyl-CoA to aldehyde, and then to ethanol by alcohol dehydrogenase.