Purification, characterization and cloning of Vibrio parahaemolyticus chitinolytic enzymes and application to oligosaccharide production

Chitinase (Pa-Chi) and chitin oligosaccharide deacetylase (Pa-COD) are involved in the production of a heterodisaccharide, beta-D-N-acetylglucosaminyl-(1,4)-D-glucosamine (GlcNAc-GlcN). These enzymes were recovered from the supernatant of Vibrio parahaemolyticus KN1699 cell culture and purified and...

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Bibliographic Details
Published in:Journal of Applied Glycoscience 2008, Vol.55(2), pp.157-164
Main Authors: Kadokura, K.(Nihon Univ., Fujisawa, Kanagawa (Japan). Coll. of Bioresource Sciences), Sakamoto, Y, Rokutani, A, Ikegami, T, Hirano, T, Yamamoto, M, Saito, K, Hakamata, W, Itoi, S, Sugita, H, Oku, T, Nishio, T
Format: Article
Language:English
Subjects:
CHEMICOPHYSICAL PROPERTIES
CHITIN
chitin oligosaccharide deacetylase
chitinase
CHITINE
CLONACION MOLECULAR
CLONAGE MOLECULAIRE
DEGRADACION
DEGRADATION
ENZIMAS
ENZYME
ENZYMES
EXPRESION GENICA
EXPRESSION DES GENES
GENE EXPRESSION
heterodisaccharide
MOLECULAR CLONING
OLIGOSACARIDOS
OLIGOSACCHARIDE
OLIGOSACCHARIDES
PRODUCCION
PRODUCTION
PROPIEDADES FISICOQUIMICAS
PROPRIETE PHYSICOCHIMIQUE
PROTEINAS RECOMBINANTES
PROTEINE RECOMBINANTE
PURIFICACION
PURIFICATION
QUITINA
recombinant enzyme
RECOMBINANT PROTEINS
Vibrio parahaemolyticus
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Summary:Chitinase (Pa-Chi) and chitin oligosaccharide deacetylase (Pa-COD) are involved in the production of a heterodisaccharide, beta-D-N-acetylglucosaminyl-(1,4)-D-glucosamine (GlcNAc-GlcN). These enzymes were recovered from the supernatant of Vibrio parahaemolyticus KN1699 cell culture and purified and characterized. For each enzyme, an ORF encoding gene and its signal sequence were cloned from genomic DNA of strain KN1699. In addition, the expression plasmid was constructed for each enzyme gene and inserted into Escherichia coli cells, and recombinant Pa-Chi and Pa-COD (Pa-rChi and Pa-rCOD) were secreted into the culture medium with the aid of signal peptides. Di-N-acetylchitobiose [(GlcNAc)sub(2)] was produced in 60% (w/w) yield by cultivating the Pa-rChi-secreting E. coli cells in 2% (w/v) beta-chitin-containing medium. Moreover, GlcNAc-GlcN was produced in high yield by treating (GlcNAc)sub(2) with the culture supernatant of Pa-rCOD-secreting E. coli cells.
ISSN:1344-7882
1880-7291
DOI:10.5458/jag.55.157