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ISSN 0582-9879 Acta Biochim et Biophysica Sinica 2004, 36(4):303-308 CN 31-1300/Q
Molecular
Cloning and Characterization of cDNA Encoding Fibrinolytic
Enzyme-3 from Earthworm Eisenia foetida
Guo-Qing DONG, Xiao-Ling
YUAN, Ya-Jun SHAN, Zhen-Hu ZHAO, Jia-Pei CHEN, and Yu-Wen CONG*
( Department of Experimental Therapy of ARS, Beijing Institute of Radiation
Medicine, Beijing 100850, China )
Abstract 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.
Key words earthworm; Eisenia
foetida; earthworm fibrinolytic enzyme; cloning; expression; refolding
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Received: December 22, 2003 Accepted:
February 19, 2004
*Corresponding author: Tel, 86-10-66930284; E-mail, [email protected]