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Gene expression and characteristics of a novel fibrinolytic enzyme (subtilisin DFE) in Escherichia coli
Author(s) -
Zhang R.H.,
Xiao L.,
Peng Y.,
Wang H.Y.,
Bai F.,
Zhang Y.Z.
Publication year - 2005
Publication title -
letters in applied microbiology
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.698
H-Index - 110
eISSN - 1472-765X
pISSN - 0266-8254
DOI - 10.1111/j.1472-765x.2005.01715.x
Subject(s) - subtilisin , escherichia coli , nattokinase , recombinant dna , bacillus amyloliquefaciens , biology , enzyme , biochemistry , expression vector , plasmid , microbiology and biotechnology , gene , fermentation
Aims: The objective of this study is to actively express a novel fibrinolytic enzyme, subtilisin DFE (douchi fibrinolytic enzyme), in Escherichia coli . Methods and Results: The DNA fragments encoding pro‐subtilisin DFE was amplified and cloned into the vector pET32a to obtain N‐terminal Trx fusion expression plasmid. The recombinant subtilisin DFE was successfully expressed and processed in the soluble fraction of E. coli BL21(DE3) in a similar fashion as the endogenous one of Bacillus amyloliquefaciens DC‐4, resulting in an active enzyme. Moreover, active enzyme can also be refolded from inclusion body. Conclusions: Active subtilisin DFE can be expressed and processed in E. coli. Significance and Impact of the Study: This study provides evidences that subtilisin DFE can be actively expressed in E. coli and the pro‐peptide is essential for guiding the proper folding into the active conformation. As such, large quantities of recombinant subtilisin DFE can be produced for pharmacological and clinical research.