葡萄糖异构酶产生菌株的筛选、发酵条件的优化及其粗酶热稳定性研究  

Screening of Strains Producing Glucose Isomerase,Optimization of Fermentation Medium and Thermal Stability of Its Crude Enzyme

作　　者：方然 汪莹莹 陈博 崔荣锦 程可馨 王馨雨 韩春铮 刘俊豪 董若云 朱圣 乔洁 曾化伟[1] FANG Ran;WANG Yingying;CHEN Bo;CUI Rongjin;CHENG Kexin;WANG Xinyu;HAN Chunzheng;LIU Junhao;DONG Ruoyun;ZHU Shen;QIAO Jie;ZENG Huawei(Anhui Provincial Key Laboratory of Pollutant-Sensitive Materials and Environment,School of Life Sciences,Huaibei Normal University,Huaibei 235000,China)

机构地区：[1]淮北师范大学生命科学学院安徽省污染物敏感材料与环境重点实验室,安徽淮北235000

出　　处：《中央民族大学学报（自然科学版）》2024年第2期16-25,共10页Journal of Minzu University of China(Natural Sciences Edition)

基　　金：国家级大学生创新创业训练计划(202210373018);安徽省科技厅重大科技专项项目(202103a06020004);安徽省大学生创新创业训练计划(202310373066,202310373067,202310373069)。

摘　　要：葡萄糖异构酶(Glucose Isomerase,GI)可以将D-葡萄糖等醛糖异构化为相应的酮糖,是高果糖浆和燃料乙醇的工业化生产中的关键酶。本实验旨在从高温大曲中筛选产葡萄糖异构酶的菌株并研究葡萄糖异构酶的热稳定性,同时,优化其液体发酵培养基。以高温大曲为筛菌样本,在细菌和真菌的初筛培养基上进行梯度稀释和平板划线,初步筛选出产葡萄糖异构酶的6株细菌和8株真菌,接着对细菌和真菌中葡萄糖异构酶酶活力高的6株菌进行热稳定性研究。选取酶活力高且热稳定性好的菌株(XJ-3)进行碳源及碳源浓度、氮源与氮源浓度,表面活性剂及表面活性剂浓度的优化,最后进行正交试验,考察培养基不同成分及浓度对细菌液体发酵葡萄糖异构酶的最优组合。结果表明:最佳培养基为:D-木糖5g/L、酵母浸出粉35 g/L、TritonX-1000.2%、MgSO_(4)·7H_(2)O 1 g/L、NaCl 10 g/L、K_(2)HPO_(4)1 g/L,预测在此组合下,葡萄糖异构酶有最大酶活404.034 U/mL,与初筛时相比,酶活提高了1.53倍。Glucose isomerase can isomerize aldose such as D-Glucose,D-Xylose to ketose,which is a key enzyme in the industrial production of high fruit syrup and fuel ethanol.This study aimed to screen glucose isomerase strains from high temperature Daqu and study the thermal stability and optimize the liquid fermentation medium.Method:Taking high temperature Daqu as the screening sample,gradient dilution and plate streaking were performed on the primary screening medium of bacteria and fungi.Six bacterial strains and 8 fungi producing glucose isomerase were selected.Then,thermal stability was performed on six strains with high glucose isomerase enzyme activity in bacteria and fungi.XJ-3 with the property of high enzyme activity and great thermal stability were selected as the strains to study on optimization carbon sources and the concentration of carbon sources,nitrogen sources and the concentration of nitrogen sources,surfactant and the concentration of surfactant.Finally,orthogonal test was performed to investigate the optimal combination of different components and concentrations of culture medium.The results showed that the optimum medium was D-xylose 5 g/L,yeast leaching powder 35 g/L,Triton X-1000.2%,MgSO_(4)·7H_(2)O 1 g/L,NaCl 10 g/L,K,HPO 1 g/L.It was predicted that the glucose isomerase has a maximum enzymatic activity of 404.034 U/mL under this combination.Compared with the initial screening period,the enzymatic activity of glucose isomerase was increased by 1.53-fold.

关 键 词：高温大曲 葡萄糖异构酶 热稳定性 液体发酵优化 正交试验 

分 类 号：Q558[生物学—生物化学]