重组木聚糖酶的高密度发酵及其定向制备低聚木糖  

High Density Fermentation of Recombinant Xylanase and Its Directional Preparation of Xylooligosaccharides

作　　者：赵剑[1] 解静聪 姜越 杨静[1] 徐浩[1] 张宁[1] ZHAO Jian;XIE Jingcong;JIANG Yue;YANG Jing;XU Hao;ZHANG Ning(Institute of Chemical Industry of Forest Products,CAF,Key Lab.of Biomass Energy and Material,Jiangsu Province,Key Lab.of Chemical Engineering of Forest Products,National Forestry and Grassland Administration,National Engineering Research Center of Low-Carbon Processing and Utilization of Forest Biomass,Jiangsu Co-Innovation Center of Efficient Processing and Utilization of Forest Resources,Nanjing 210042,China)

机构地区：[1]中国林业科学研究院林产化学工业研究所、江苏省生物质能源与材料重点实验室、国家林业和草原局林产化学工程重点实验室、林木生物质低碳高效利用国家工程研究中心、江苏省林业资源高效加工利用协同创新中心,江苏南京210042

出　　处：《林产化学与工业》2024年第1期97-104,共8页Chemistry and Industry of Forest Products

基　　金：国家重点研发计划资助项目(2019YFB1503803);江苏省自然科学基金青年项目(BK20180153)。

摘　　要：为提高酶水解杨木木聚糖定向转化低聚木糖的效率,同时降低酶法制备低聚木糖的成本,围绕重组大肠杆菌(PET-PdoXyn10A-DE3)高密度发酵制备拟杆菌来源木聚糖酶PdoXyn10A的诱导表达条件和酶水解制备低聚木糖的工艺条件开展了研究。考察了在5 L发酵罐水平下诱导剂浓度、诱导光密度(OD_(600))值、诱导温度和诱导pH值等因素对PET-PdoXyn10A-DE3产重组木聚糖酶PdoXyn10A的影响,反应温度、pH值对玉米芯木聚糖酶水解过程中酶活力的影响,以及酶添加量、酶水解时间对杨木乙酸水解液制备低聚木糖的影响。研究结果表明:在诱导剂浓度0.25 mmol/L、诱导OD_(600)值55、诱导温度34℃和诱导pH值7.0的最佳诱导条件下,经高密度发酵重组酶酶活力可达362.67 U/mL,重组大肠杆菌生物量可达28.83 g/L。酶水解制备低聚木糖的最佳条件为酶反应温度40℃、pH值5.2、酶添加量12 U/mL和反应时间6 h,此条件下低聚木糖的量为3.21 g/L,其中木二糖、木三糖、木四糖、木五糖和木六糖的量分别为1.09、 0.97、 0.84、 0.18、 0.13 g/L,木糖为1.82 g/L。In order to improve the enzymatic hydrolysis efficiency of poplar xylan to xylo-oligosaccharides and reduce the cost of enzymatic preparation of xylo-oligosaccharides,this study focused on the induced expression conditions of Bacteroides-derived xylanase PdoXyn10A produced by high-density fermentation of recombinant Escherichia coli(PET-PdoXyn10A-DE3) and the process conditions of enzymatic hydrolysis to prepare xylo-oligosaccharides.The effects of the inducer concentration,the induced optical density(OD_(600)) value,induction temperature and induction pH value on the production of recombinant xylanase PdoXyn10A by PET-PdoXyn10A-DE3 were studied under 5 L fermenter level.The effects of reaction temperature and pH value on the enzyme activity of corncob xylanase during hydrolysis and the effects of enzyme addition and hydrolysis time on the preparation of xylo-oligosaccharide from poplar acetic acid hydrolysate were determined.The results showed that the activity of recombinase reached 362.67 U/mL and the biomass of recombinant Escherichia coli reached 28.83 g/L under high density fermentation at the optimal induction conditions with 0.25 mmol/L of inducer concentration,induction OD_(600) value of 55,induction temperature 34 ℃ and induction pH value 7.0.The optimum conditions for the preparation of xylooligosaccharides by enzymatic hydrolysis were as follows:enzyme reaction temperature of 40 ℃,pH value of 5.2,the enzyme dosage of 12 U/mL,and reaction time of 6 h.Under these conditions,the content of xylo-oligosaccharides was 3.21 g/L,the content of xylobiose,xylotriose,xylotetraose,xylopentaose,xylohexaose were 1.09,0.97,0.84,0.18,0.13 g/L,respectively,and the content of xylose was 1.82 g/L.

关 键 词：重组木聚糖酶 高密度发酵 乙酸水热反应 低聚木糖 酶水解条件 

分 类 号：TQ35[化学工程]