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Efficient Expression and Purification of Recombinant Human Enteropeptidase Light Chain in Esherichia coli BABT
Niu,Li-Xi; Li,Jia-Yue; Ji,Xue-Xue; Yang,Bin-Sheng.
Human enterokinase (synonym: enteropeptidase, EC 3.4.21.9) light chain (hEKL) gene was designed and artificially synthesized with built-in codon blas towards Escherichia colicodon preference. The synthetic hEKL gene was cloned into prokaryotic expression vector pMAL-s and transferred into the expression strain E. coli BL21 (DE3). Recombinant hEKL protein with a maltose binding protein (MBP) tag was expressed at high levels in soluble form, which yielded about 42% of the total cellular protein. The target protein was then purified to the homogeneity (> 95%) by affinity chromatography. The peptide substrate GST-Melittin with enterokinase recognition site was completely cleaved by the purified MBP-hEKL at the molar ratio of 1:5000 (enzyme:substrate)....
Tipo: Info:eu-repo/semantics/article Palavras-chave: Human enterokinase light chain (hEKL); Fusion expression; Affinity purification; Activity analysis; Enzymatic cleavage.
Ano: 2015 URL: http://www.scielo.br/scielo.php?script=sci_arttext&pid=S1516-89132015000200154
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