Molecular cloning and characterization of cDNA encoding fibrinolytic enzyme-3 from earthworm Eisenia foetida.
The earthworm fibrinolytic enzyme-3 (EFE-3, GenBank accession No: AY438622), from the earthworm Eisenia foetida, is a component of earthworm fibrinolytic enzymes. In this study, cDNA encoding the EFE-3 was cloned by RT-PCR. The cDNA contained an open reading frame of 741 nucleotides, which encoded a deduced protein of 247 amino acid residues, including signal sequences. EFE-3 showed a high degree of homology to earthworm (Lumbricus rebullus) proteases F-III-1, F-III-2, and bovine trypsin. The recombinant EFE-3 was expressed in E. coli as inclusion bodies, and the gene encoding the native form of EFE-3 was expressed in COS-7 cells in the medium. Both the refolding product of inclusion bodies and the secreted protease could dissolve the artificial fibrin plate.
Dong
Guo-Qing
GQ
Department of Experimental Therapy of ARS, Beijing Institute of Radiation Medicine, China.
Yuan
Xiao-Ling
XL
Shan
Ya-Jun
YJ
Zhao
Zhen-Hu
ZH
Chen
Jia-Pei
JP
Cong
Yu-Wen
YW
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D019556:COS Cells; D002522:Chlorocebus aethiops; D003001:Cloning, Molecular; D019295:Computational Biology; D003470:Culture Media; D018076:DNA, Complementary / Q000235:genetics*; D004591:Electrophoresis, Polyacrylamide Gel; D010450:Endopeptidases / Q000737:chemistry* / Q000235:genetics* / Q000378:metabolism; D004926:Escherichia coli / Q000235:genetics; D005342:Fibrinolysis; D015870:Gene Expression; D002479:Inclusion Bodies / Q000378:metabolism; D008969:Molecular Sequence Data; D009835:Oligochaeta / Q000201:enzymology* / Q000235:genetics*; D016366:Open Reading Frames; D017510:Protein Folding; D021382:Protein Sorting Signals; D011994:Recombinant Proteins / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction; D017422:Sequence Analysis, DNA; D017386:Sequence Homology, Amino Acid; D012689:Sequence Homology, Nucleic Acid
D003470:Culture Media; D018076:DNA, Complementary; D021382:Protein Sorting Signals; D011994:Recombinant Proteins; D010450:Endopeptidases; C083421:lumbrokinase