1. Molecular characterization of a cold-active recombinant xylanase from Flavobacterium johnsoniae and its applicability in xylan hydrolysis.
- Author
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Chen S, Kaufman MG, Miazgowicz KL, Bagdasarian M, and Walker ED
- Subjects
- Amino Acid Sequence, Cold Temperature, Endo-1,4-beta Xylanases genetics, Enzyme Activation, Enzyme Stability, Flavobacteriaceae classification, Hydrolysis, Molecular Sequence Data, Molecular Weight, Species Specificity, Endo-1,4-beta Xylanases chemistry, Flavobacteriaceae enzymology, Recombinant Proteins chemistry, Xylans chemistry
- Abstract
A novel xylanase gene, xyn10A, was cloned from Flavobacterium johsoniae, overexpressed in a flavobacterial expression system, the recombinant enzyme purified by Ni-affinity chromatography, and enzyme structure and activity analyzed. Xyn10A was found to be a modular xylanase with an Fn3 accessory domain on its N-terminal and a catalytic region on the C-terminal. The optimum pH and temperature for Xyn10A was 8.0 and 30 °C, but Xyn10A retained 50% activity at 4 °C, indicating that Xyn10A is a cold-active xylanase. A Fn3-deletion xylanase had relative activity ca. 3.6-fold lower than the wild-type, indicating that Fn3 promotes xylanase activity. The Fn3 region also contributed to stability of the enzyme at elevated temperatures. However, Fn3 did not bind this xylanase to insoluble substrates. The enzyme hydrolyzed xylo-oligosaccharides into xylobiose, and xylose with xylobiose as the main product, confirming that Xyn10A is a strict endo-β-1,4-xylanase. Xyn10A also hydrolyzed birchwood and beechwood xylan to yield mainly xylose, xylobiose and xylotriose., (Copyright © 2012 Elsevier Ltd. All rights reserved.)
- Published
- 2013
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