Production of a recombinant chitin oligosaccharide deacetylase from Vibrio parahaemolyticus in the culture medium of Escherichia coli cells |
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Authors: | Kazunari Kadokura Yusuke Sakamoto Kaori Saito Takanori Ikegami Takako Hirano Wataru Hakamata Tadatake Oku Toshiyuki Nishio |
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Institution: | (1) Department of Biological Chemistry, College of Bioresource Sciences, Nihon University, 1866 Kameino, Fujisawa Kanagawa, 252-8510, Japan |
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Abstract: | An open reading frame (ORF) encoding chitin oligosaccharide deacetylase (Pa-COD) gene and its signal sequence was cloned from the Vibrio parahaemolyticus KN1699 genome and its sequence was analyzed. The ORF encoded a 427 amino acid protein, including the 22 amino acid signal
sequence. The deduced amino acid sequence was highly similar to several bacterial chitin oligosaccharide deacetylases in carbohydrate
esterase family 4. An expression plasmid containing the gene was constructed and inserted into Escherichia coli cells and the recombinant enzyme was secreted into the culture medium with the aid of the signal peptide. The concentration
of the recombinant enzyme in the E. coli culture medium was 150 times larger than that of wild-type enzyme produced in the culture medium by V. parahaemolyticus KN1699. The recombinant enzyme was purified to homogeneity from culture supernatant in an overall yield of 16%. Substrate
specificities of the wild-type and the recombinant enzymes were comparable. |
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Keywords: | Chitin oligosaccharide deacetylase Escherichia coli Recombinant enzyme Signal peptide Vibrio parahaemolyticus |
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