β-琼脂糖酶(AgaA7)在大肠杆菌中的表达、纯化及酶学性质研究  

Expression,Purification and Enzymatic Characterization ofβ-Agarase(AgaA7) in Escherichia coli

作　　者：王广慧[1] 

机构地区：[1]绥化学院化学与制药工程系,黑龙江绥化152061

出　　处：《佳木斯大学学报（自然科学版）》2008年第3期416-420,共5页Journal of Jiamusi University：Natural Science Edition

基　　金：黑龙江省教育厅科学技术研究项目(项目编号:11511465)

摘　　要：为研究重组β-琼脂糖酶（AgaA7）基因的表达产物及其酶学性质,根据GeneBank中β-琼脂糖酶（AgaA7）基因的编码序列合成此基因后,将其克隆到表达载体pET-22b（＋）中,并在大肠杆菌BL21（DE3）中进行表达.纯化后的表达产物经SDS-PAGE检测其相对分子质量并测定其酶学性质.结果表明：重组酶的相对分子质量为～5.0×10^4,与预期相符.此酶最适反应pH是7.0,在pH58的缓冲液中最稳定;最适反应温度为50℃,在低于50℃时比较稳定.Na^＋,K^＋,Ca^2＋,Mg^2＋对重组酶活性影响不大,而Zn^2＋,Hg^2＋,Pb^2＋则对酶活性具有不可逆抑制作用.In order to study the expression product of recombinantβ-agarase（AgaA7） gene and its enzymatic characterization,the full β-agarase（AgaA7） sequence is synthesized according to the Gene Bank.Then,it is inserted into expression vector pET-22b（＋）.The recombinant plasmid is transformed into E.coli BL21（DE3）.The molecular mass of the purified expression product is examined by SDS-PAGE and its bioactivities are proved.The optimum reactive pH and temperature of the recombinantβ-agarase are 7.0 and 50℃ respectively.The enzyme is most stable at pH58 and below 50℃.Na^＋,K^＋,Ca^2＋ and Mg^2＋ ions don＇t affect it＇s activity,but Zn^2＋,Hg^2＋ and Pb^2＋ ions inactivate it irreversibly.

关 键 词：β-琼脂糖酶(AgaA7) 表达 纯化 酶学性质 

分 类 号：Q936[生物学—微生物学] Q78