Abstract
Two cDNA fragments (lrF1 and lrF2) representing a fibrinolytic enzyme gene of F-III-2 (GenBank AB045719), without and with signal peptide coding sequence, were cloned from earthworm Lumbricus rubellus. The two fragments were inserted into bacterial expression vector pET28a (+), respectively. Subsequent expression showed that both lrF1 and lrF2 proteins were produced as an inclusion body form in E. coli BL21 (DE3) pLysE. After protein refolding and purification, the fusion lrF1 and its derivative without poly histidine tags at the N-terminus showed fibrinolytic activity on fibrin plates with relative activity of 134.3 U/mg protein and 139.7 U/mg protein, respectively, whereas the fusion lrF2 and its derivative without the tags at the N-terminus, had no fibrinolytic activity. The results indicated that the E. coli expression system could not recognize the endogenous signal peptide of F-III-2, and the effect of the histidine tags at the N-terminus on the fibrinolytic activity of the expressed protein was insignificant.
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Li, Dh., Tong, W. & Yang, Yf. Functional expression of an earthworm fibrinolytic enzyme in Escherichia coli . World J Microbiol Biotechnol 24, 613–618 (2008). https://doi.org/10.1007/s11274-007-9515-3
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DOI: https://doi.org/10.1007/s11274-007-9515-3