普鲁兰酶N端结构域CBM68对酶学性质的影响  

Effect of the N-terminal Domain CBM68 on the Enzymatic Properties of Pullulanase

作　　者：曾艳 郑宏臣[2,3] 付晓平[2,3] 徐健勇[2,3] 宋诙[2,3] 刘方[1] Zeng Yan;Zheng Hongchen;Fu Xiaoping;Xu Jianyong;Song hui;Liu Fang(College of Life Science,Nankai University,Tianjin 300071,China;Industrial Enzymes National Engineering Laboratory,Tianjin Institute of Industrial Biotechnology,Chinese Academy of Sciences,Tianjin 300308,China;Key Laboratory of Tianjin Industrial Biosystem and Process Engineering,Tianjin Institute of Industrial Biotechnology,Chinese Academy of Sciences,Tianjin 300308,China)

机构地区：[1]南开大学生命科学学院,天津300071 [2]中国科学院天津工业生物技术研究所工业酶国家工程实验室,天津300308 [3]中国科学院天津工业生物技术研究所天津工业生物系统与过程重点实验室,天津300308

出　　处：《南开大学学报（自然科学版）》2018年第4期93-99,共7页Acta Scientiarum Naturalium Universitatis Nankaiensis

基　　金：国家自然科学基金(31701534);中国科学院重点部署项目(kfj-sts-zdtp-016-1;kfzd-sw-211-2);天津市科技计划项目(16yfzcsy00790;16yfxtsy00530;15yfyssy00040)

摘　　要：碳水化合物结合模块(Carbohydrate Binding Modules,CBMs)在普鲁兰酶结构中普遍存在,对酶的催化性质起重要作用.本文将酸性普鲁兰酶Pul B的N端结构域CBM41-X45替换成耐热普鲁兰酶Pul A的N端结构域CBM68,得到重组酶Pul-11,同时将Pul-11的催化结构域替换成Pul A的催化结构域,得到重组酶Pul-12.结果显示,重组酶Pul-11和Pul-12的最适温度均较Pul B提高了10℃,最适pH分别提高了0.5和1;60℃下热稳定性分别提高了41.4%和44.0%;同时,重组酶Pul-11和Pul-12的底物亲和力和催化效率较Pul B也均有一定程度提高.可见,来源于耐热普鲁兰酶的新型底物结构域CBM68对普鲁兰酶的最适作用温度、最适作用pH及催化效率具有重要影响.Carbohydrate Binding Modules（CBMs） are common domains in pullulanase, which play important roles on the catalytic properties of glycosyl hydrolases. In this work, the N-terminal domain CBM41-X45 of the acid pullulanase Pul B was replaced by CBM68, which is the N-terminal domain of the thermostable pullulanase Pul A. And then the catalytic domain of the recombinant pullulanase was further replaced by the catalytic domain of Pul A. The two recombinant pullulanases were named as Pul-11 and Pul-12, respectively. The same optimum temperature of Pul-11 and Pul-12 was 65 ℃, which was10 ℃ higher than that of Pul B. And the optimum pH of Pul-11 and Pul-12 were higher than that of Pul B by 0.5 and 1 unit, respectively. The residual activities of Pul-11 and Pul-12 were 41.4 % and44.0 % higher than that of Pul B at 60 ℃ for 12 h. Moreover, Pul-11 and Pul-12 also showed higher in substrate affinity and catalytic efficiency than Pul B. Therefore, the new substrate combination domain CBM68 shows important effects on the optimum temperature, the optimum pH and catalytic efficiency of the pullulanases through this study.

关 键 词：碳水化合物结合模块 CBM68 普鲁兰酶 最适作用温度 最适作用pH 催化效率 

分 类 号：Q814[生物学—生物工程]