两种不同固定化方法对木聚糖酶Xyn11A酶学性质的影响及在低聚木糖制备中的应用  

Effects of two different immobilization methods on enzymatic properties of xylanase Xyn11A and applications in production of xylooligosaccharides

作　　者：李婵娟 汪松波[1] 吴高兵 LI Chanjuan;WANG Songbo;WU Gaobing(College of Food and Biotechnology,Wuhan Institute of Design and Sciences,Wuhan 430205,China;College of Life Science and Technology,Huazhong Agricultural University,Wuhan 430070,China;College of Plant Science and Technology,Huazhong Agricultural University,Wuhan 430070,China)

机构地区：[1]武汉设计工程学院食品与生物科技学院,湖北武汉430205 [2]华中农业大学生命科学技术学院,湖北武汉430070 [3]华中农业大学植物科学技术学院,湖北武汉430070

出　　处：《食品与发酵工业》2024年第10期17-23,共7页Food and Fermentation Industries

基　　金：国家自然科学基金青年科学基金项目(32101895);湖北省自然科学基金项目(2022CFB567);湖北省教育厅科学研究计划指导性项目(B2022424);武汉设计工程学院校级科学研究项目(K202112)。

摘　　要：为了提高木聚糖酶Xyn11A的稳定性及在低聚木糖生产中的应用潜力,该研究利用有机-无机杂化纳米花技术和以氨基乙基-琼脂糖为载体共价结合法固定Xyn11A,制备了固定化酶Xyn11A-Cu_(3)(PO_(4))_(2)和Xyn11A-Agarose,并比较研究了2种固定化酶的酶学性质,发现2种固定化酶的pH稳定性和热稳定性均有所提高,Xyn11A-Agarose稳定性更佳,在pH 4.0~4.5处理24 h后仍有40%~65%的残余活性,50℃处理3 h后仍有60%的残余活性。以榉木木聚糖为底物,与游离酶相比,Xyn11A-Cu_(3)(PO_(4))_(2)和Xyn11A-Agarose对底物的亲和力和催化效率有所降低,但有较好的重复使用次数。特别是Xyn11A-Agarose循环使用12轮后,仍有90%的残余活性。Xyn11A-Cu_(3)(PO_(4))_(2)水解甘蔗渣和玉米芯粉生成的低聚木糖中木二糖和木三糖含量最高,木四糖含量最低;而Xyn11A-Agarose催化生成的不同低聚木糖含量相当。2种固定化方法均能显著提高Xyn11A的稳定性,但Xyn11A-Agarose的稳定性和重复性优于Xyn11A-Cu_(3)(PO_(4))_(2),表明Xyn11A-Agarose在生产低聚木糖中具有更大应用潜力。In this study,the immobilization of xylanase Xyn11A using a protein-inorganic hybrid nanoflower system and covalent immobilization with aminoethyl-agarose were assessed to improve the enzyme properties.The enzymatic properties of Xyn11A-Cu_(3)(PO_(4))2 and Xyn11A-Agarose were compared.It was found that the pH stability and thermal stability of the two immobilized enzymes were improved,and Xyn11A-Agarose was more stable.Xyn11A-Agarose retained 40%-65%residual activity after incubation at pH 4.0-4.5 buffer for 24 h and 60%residual activity at 50℃for 3 h,respectively.Xyn11A-Cu_(3)(PO_(4))2 and Xyn11A-Agarose hydrolyzed beechwood xylan with K m of(10.14±3.56)mg/mL and(21.52±2.33)mg/mL,2.39 and 5.06 times of free Xyn11A,respectively.The K cat of Xyn11A-Cu_(3)(PO_(4))_(2)was 1.87 times higher than that of Xyn11A,but the K cat/K m decreased by about 21%.The K cat of Xyn11A-Agarose was similar to the free enzyme,but the K cat/K m was only 22%of the free enzyme.After 6 cycles of reuse,Xyn11A-Cu_(3)(PO_(4))_(2)retained 30%residual activity,while Xyn11A-Agarose still presented 90%residual activity after 12 cycles of reuse.The xylooligosaccharides produced by the hydrolysis of bagasse and corncob flour by Xyn11A-Cu_(3)(PO_(4))_(2)had the highest content of xylobiose and xylotriose,and the lowest content of xylotetraose,while the different xylooligosaccharides catalyzed by Xyn11A-Agarose were almost equivalent in content.The stability and reusability of Xyn11A-Agarose were better than those of Xyn11A-Cu_(3)(PO_(4))_(2),indicating that Xyn11A-Agarose has greater application potential in the production of xylooligosaccharides.

关 键 词：木聚糖酶 固定化 酶学性质 低聚木糖 

分 类 号：O636.1[理学—高分子化学]