产甘油假丝酵母补料发酵中的甘油合成衰减

Glycerol Synthesis Attenuation of Candida glycerinogenes in Fed-batch Fermentation

为揭示产甘油假丝酵母Candida glycerinogenes补料发酵后甘油合成衰减机理,以指导甘油合成及菌株改造,对补料发酵后甘油的生成情况、葡萄糖的消耗情况、细胞内的代谢流量变化以及代谢途径关键酶活力变化进行了研究.结果表明,补料发酵后葡萄糖的代谢流量分布发生变化,流向HMP途径和甘油合成途径的流量分别降低48%和33%,更多的碳流通过EMP途径形成副产物;胞浆3-磷酸甘油脱氢酶(ctGPD)和6-磷酸葡萄糖脱氢酶(G6PDH)的活力下降,而丙酮酸激酶(PYK)活力上升.补料发酵后甘油合成关键酶ctGPD酶活力低下,流向甘油合成途径的碳流减小,还原力供给受影响,这些因素共同作用引起了补料发酵后甘油合成的衰减.

The previous resulted showed that in fed-batch fermentation of Candida glycerinogenes,the glycerol concentration decreased by 13%,the conversion ratio of glycerol decreased by 34%,the glycerol synthesis rate decreased by 67% and the glucose consumption rate decreased by 44%.In order to reveal the mechanism of glycerol synthesis attenuation,the glycerol synthesis,glucose consumption,intracellular metabolic flux distribution and the activity of key enzymes in metabolism were investigated.It was found that the metabolic flux distribution changed compared with batch fermentation.The metabolic flux decreased by 48% and 33% respectively in HMP and glycerol synthesis pathway while the metabolic flux increased in EMP,which led to the increase of by-products.The activity of ctGPD and G6PDH decreased,while the activity of PYK increased.ctGPD,a key enzyme for glycerol formation,directly affected the glycerol formation.G6PDH influenced the carbon flux to HMP,which not only played an important role in cell growth,but also provided reducing power for glycerol formation.PYK influenced the metabolic flux distribution between EMP and glycerol synthesis pathway.Consequently,the decrease in the activity of G6PDH and carbon flux to glycerol synthesis pathway,and inefficient reducing power for glycerol formation showed synergy effects on the glycerol synthesis attenuation in fed-batch fermentation.The results should provide the primary datamaterials for the regulation of glycerol synthesis in C.glycerinogenes and strain improvement.