Expression and characterization of soluble recombinant Ulp1p with glutathione S-transferase tag in Escherichia coli

The aim of the study is to obtain an efficient expression of recombinant ubiquitin-like specific protease 1 (Ulp1) by gene engineering. We cloned the Ulp1p, active fragment (403 aa-621 aa) of Ulp1, from Saccharomyces cerevisia, and subcloned into pGEX/Rosetta (DE3) to form an expression plasmid, pGE...

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Published inShengwu gongcheng xuebao Vol. 26; no. 6; p. 837
Main Authors Fu, Junhua, Wang, Qi, Yin, Jiechao, Liu, Mingyao, Li, Ning, Yao, Wenbin, Ren, Guiping, Li, Lu, Li, Deshan
Format Journal Article
LanguageChinese
Published China 01.06.2010
Subjects
Cloning, Molecular
Cysteine Endopeptidases - biosynthesis
Cysteine Endopeptidases - genetics
Escherichia coli - genetics
Escherichia coli - metabolism
Fungal Proteins - biosynthesis
Fungal Proteins - genetics
Glutathione Transferase - biosynthesis
Glutathione Transferase - genetics
Recombinant Fusion Proteins - biosynthesis
Recombinant Fusion Proteins - genetics
Saccharomyces cerevisiae - enzymology
Solubility
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Summary:The aim of the study is to obtain an efficient expression of recombinant ubiquitin-like specific protease 1 (Ulp1) by gene engineering. We cloned the Ulp1p, active fragment (403 aa-621 aa) of Ulp1, from Saccharomyces cerevisia, and subcloned into pGEX/Rosetta (DE3) to form an expression plasmid, pGEX-Ulp1p-His6. In order to enhance the solubility of GST-Ulp1p-His6, we purified the fusion protein GST-Ulp1p-His6 by either glutathione S-transferase agarose or Ni-NTA resin chromatography, the purity was up to 98%. We utilized the protein to cleave the SUMO fusions, and the specific activity of GST-Ulp1p-His6 was 1.375 x 10(4) U/mg. This study showed that the recombinant protein GST-Ulp1p-His6 displayed high specificity and activity.
ISSN:1000-3061