cDNA cloning and expression of earthworm fibrinolytic enzyme component A  

作　　者：LIUJunfeng WANGXinquan 等 

机构地区：[1]NationalLaboratoryofBiomacromolecules,InstituteofBiophysics,ChineseAcademyofSciences,Beijing100101,China [2]NationalLaboratoryofBiomacromolecules,InstituteofBiophysics,ChineseAcademyofSciences,Beijing100

出　　处：《Chinese Science Bulletin》2003年第1期68-71,共4页

基　　金：This work was supported by the National Natural Science Foundation of China (Grant No. 39970174) and the Foundation of the National Key Research Development Project of China (Grant No. G1999075601).

摘　　要：Earthworm fibrinolytic enzyme component A (EFEa), a protein with dual fibrinolytic activity, is one of the major therapeutically important earthworm fibrinolytic enzyme components. The cDNA fragment encoded the ma-ture protein was cloned from earthworm (Eisenia fetida) by the RT-PCR technique. The deduced amino acid sequence of the EFE component A shows high homology with some members of serine proteases trypsin family, and the amino acid residues constituting the active sites are conserved in the EFEa as compared with the other proteins of the trypsin family. The cDNA fragment was subcloned into the expres-sion vector pQE31 and pMAL-c2X of E. coli. The resulting expression plasmids, pQE-efea and pMAL-efea, were used to transform the E. coli strain M15. Recombinant protein bands corresponding with calculated molecular weights were induced. The induced His6-EFEa fusion protein with pQE- efea was accumulated into inclusion body, while the induced MBP-EFEa fusion protein with pMAL-efea was soluble and showed fibrinolytic activities.Earthworm fibrinolytic enzyme component A (EFEa), a protein with dual fibrinolytic activity, is one of the major therapeutically important earthworm fibrinolytic enzyme components. The cDNA fragment encoded the ma-ture protein was cloned from earthworm (Eisenia fetida) by the RT-PCR technique. The deduced amino acid sequence of the EFE component A shows high homology with some members of serine proteases trypsin family, and the amino acid residues constituting the active sites are conserved in the EFEa as compared with the other proteins of the trypsin family. The cDNA fragment was subcloned into the expres-sion vector pQE31 and pMAL-c2X of E. coli. The resulting expression plasmids, pQE-efea and pMAL-efea, were used to transform the E. coli strain M15. Recombinant protein bands corresponding with calculated molecular weights were induced. The induced His6-EFEa fusion protein with pQE- efea was accumulated into inclusion body, while the induced MBP-EFEa fusion protein with pMAL-efea was soluble and showed fibrinolytic activities.

关 键 词：蚯蚓 纤溶酶成分A CDNA 基因克隆 基因表达 

分 类 号：Q789[生物学—分子生物学] R973.2[医药卫生—药品]