人参皂苷β-葡萄糖苷酶基因的毕赤酵母载体构建及生物转化  

Construction of Ginsenoside-β-glucosidase Gene Vector and Biotransformation in Pichia Pastoris

作　　者：刘欣茹 刘春莹 徐龙权[1] 宋建国 鱼红闪[1] LIU Xinru;LIU Chunying;XU Longquan;SONG Jianguo;YU Hongshan(School of Biology Engineering,Dalian Polytechnic University,Dalian 116034,China;School of Life Science and Technology,Dalian University,Dalian 116622,China)

机构地区：[1]大连工业大学生物工程学院,大连116034 [2]大连大学生命科学与技术学院,大连116622

出　　处：《高等学校化学学报》2018年第11期2451-2457,共7页Chemical Journal of Chinese Universities

基　　金：国家高端外国专家项目(批准号:GDT20152100019)资助~~

摘　　要：将人参皂苷β-葡萄糖苷酶(Glu GF)基因与表达载体p PIC9K连接,构建重组质粒,转化至毕赤酵母中表达,并用于人参皂苷Rb1的催化转化.研究结果表明,成功构建了重组表达质粒p PIC9K-GluGF,转化后筛选到阳性重组毕赤酵母菌.重组菌产酶的最佳甲醇诱导体积为0. 5%,诱导时间为168 h.重组菌诱导培养制备的粗酶液具有Glu GF的活性,可以水解人参皂苷Rb1,产物中皂苷C-K含量达到0. 09 mg/mL,粗酶液中Glu GF的比活力为0. 67 U/mg.The ginsenoside-β-glucosidase(GluGF)gene was ligated with the expression vector pPIC9K to construct a recombinant plasmid.The recombinant plasmid was transformed into pichia pastoris to expresse and accomplish the biotransformate of ginsenoside Rb 1.The results showed that the recombinant plasmid pPIC9K-GluGF was successfully constructed and transformed into pichia pastoris GS115 competent cells.The optimal volume of methanol for recombinant strain was 0.5%,and the induction time was 168 h.The crude enzyme solution prepared by induction culture could hydrolyze ginsenoside Rb 1.The content of C-K in the product reached 0.09 mg/mL.The enzyme activity of GluGF in the crude enzyme solution was 0.67 U/mg.

关 键 词：人参皂苷β-葡萄糖苷酶 人参皂苷RB1 载体构建 毕赤酵母 生物转化 

分 类 号：O629[理学—有机化学]