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KMID : 0613820070170091197
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Journal of Life Science
2007 Volume.17 No. 9 p.1197 ~ p.1203
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Cloning, Sequencing and Expression of the Gene Encoding a Thermostable β-Xylosidase from Paenibacillus sp. DG-22
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Lee Tae-Hyeong
Lee Yong-Eok
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Abstract
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A genomic DNA library of the bacterium Paenibacillus sp. DG-22 was constructed and the β-xylosidase-positive clones were identified using the fluorogenic substrate 4-methylumbelliferyl-β-D-xylopyranoside (βMUX). A recombinant plasmid was isolated from the clone and 4.3-kb inserted DNA was sequenced. The β-xylosidase gene (xylA) was comprised of a 2,106 bp open reading frame (ORF) encoding 701 amino acids with a molecular weight of 78,710 dalton and a pI of 5.0. The deduced amino acid sequence of the xylA gene product had significant similarity with β-xylosidases classified into family 52 of glycosyl hydrolases. The xylA gene was subcloned into the pQE60 expression vector to fuse with six histidine-tag. The recombinant β-xylosidase (XylA-H6) was purified to homogeneity by heat-treatment and immobilized metal affinity chromatography. The pH and temperature optima of the XylA-H6 enzyme were pH 5.5-6.0 and 60℃, respectively.
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KEYWORD
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Paenibacillus sp. β-xylosidase, cloning, expression
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