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解淀粉芽孢杆菌DC-4豆豉溶栓酶成熟肽编码序列的克隆及表达
引用本文:彭勇,张义正.解淀粉芽孢杆菌DC-4豆豉溶栓酶成熟肽编码序列的克隆及表达[J].应用与环境生物学报,2002,8(3):285-289.
作者姓名:彭勇  张义正
作者单位:四川大学生命科学学院分子生物学实验室,成都,610064
摘    要:利用PCR方法从解淀粉芽孢杆菌DC 4总DNA中扩增出豆豉溶栓酶 (DFE)成熟肽编码区片段 .测序结果表明 :DFE成熟肽编码区长 82 5bp,编码 2 75个氨基酸残基 ,分子量为 2 7.7× 10 3 ,推导的N’ -端氨基酸序列与豆豉溶栓酶N’ -端氨基酸测序结果完全一致 ,说明克隆到的基因确实是豆豉溶栓酶基因 .同源性分析表明 ,DFE成熟肽编码区的核苷酸和氨基酸序列与日本纳豆激酶的同源性分别为 80 .0 %和 86 .5 % ,这提示豆豉溶栓酶可能是一种新型的溶栓酶 .将表达质粒pET Nde转化E .coliBL2 1(DE3)中 ,IPTG可诱导表达大量的DFE融合蛋白 ,占菌体可溶性蛋白的 4 0 % ,主要以包涵体的形式存在 .图 3表 1参 19

关 键 词:解淀粉芽孢杆菌DC-4  豆鼓  溶栓酶  成熟肽  编码序列  克隆  表达
修稿时间:2001年12月12

CLONING AND EXPRESSION IN E. coli OF CODING SEQUENCE OF THE DOUCHI FIBRINOLYTIC ENZYME MATURE PEPTIDE FROM BACILLUS AMYLOLIQUEFACIENS DC-4
PENG Yong,&,ZHANG Yizheng.CLONING AND EXPRESSION IN E. coli OF CODING SEQUENCE OF THE DOUCHI FIBRINOLYTIC ENZYME MATURE PEPTIDE FROM BACILLUS AMYLOLIQUEFACIENS DC-4[J].Chinese Journal of Applied and Environmental Biology,2002,8(3):285-289.
Authors:PENG Yong  &  ZHANG Yizheng
Institution:PENG Yong & ZHANG Yizheng *
Abstract:The fragment encoding the douchi fibrinolytic enzyme(DFE) mature peptide was amplified from Bacillus amyloliquefaciens DC-4 total DNA by PCR. The result of sequencing showed that the coding region of the DFE mature peptide had 825 bp in length and encoded 275 amino acid residues, whose sequence was identical to the N'-terminal amino acid sequence of the purified DFE determined by Edman method, indicating that the cloned fragment was indeed as expected. Sequence homologous analysis displayed that there were 80.0% and 86.5% of nucleotide and amino acid homology between DFE and nattokinase gene, respectively, which suggested that the DFE be a novel fibrinolytic enzyme. The expression plasmid pET Nde was introduced into E.coli BL21(DE3), the His DFE fusion protein was highly expressed by IPTG induction and accumulated as inclusion body up to 40% of bacterial soluble protein. Fig 3, Tab 1, Ref 19
Keywords:douchi  Bacillus amyloliquefaciens  fibrinolytic enzyme  gene cloning  gene expression
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