Importance of the carbohydrate-binding module of Clostridium stercorarium Xyn10B to xylan hydrolysis.
The Clostridium stercorarium xylanase Xyn10B is a modular enzyme comprising two thermostabilizing domains, a family 10 catalytic domain of glycosyl hydrolases, a family 9 carbohydrate-binding module (CBM), and two S-layer homologous (SLH) domains [Biosci. Biotechnol. Biochem., 63, 1596-1604 (1999)]. To investigate the role of this CBM, we constructed two derivatives of Xyn10B and compared their hydrolytic activity toward xylan and some preparations of plant cell walls; Xyn10BdeltaCBM consists of a catalytic domain only, and Xyn10B-CBM comprises a catalytic domain and a CBM. Xyn10B-CBM bound to various insoluble polysaccharides including Avicel, acid-swollen cellulose, ball-milled chitin, Sephadex G-25, and amylose-resin. A cellulose binding assay in the presence of soluble saccharides suggested that the CBM of Xyn10B had an affinity for even monosaccharides such as glucose, galactose, xylose, mannose and ribose. Removal of the CBM from the enzyme negated its cellulose- and xylan-binding abilities and severely reduced its enzyme activity toward insoluble xylan and plant cell walls but not soluble xylan. These findings clearly indicated that the CBM of Xyn10B is important in the hydrolysis of insoluble xylan. This is the first report of a family 9 CBM with an affinity for insoluble xylan in addition to crystalline cellulose and the ability to increase hydrolytic activity toward insoluble xylan.
Faculty of Bioresources, Mie University, Tsu, Japan.
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Ali
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Hayashi
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Karita
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Goto
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Kimura
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Sakka
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Ohmiya
D002384:Catalysis; D002482:Cellulose / Q000378:metabolism; D003013:Clostridium / Q000201:enzymology* / Q000235:genetics / Q000378:metabolism; D004591:Electrophoresis, Polyacrylamide Gel; D006868:Hydrolysis; D008969:Molecular Sequence Data; D017434:Protein Structure, Tertiary; D043325:Xylan Endo-1,3-beta-Xylosidase; D014990:Xylans / Q000378:metabolism*; D014995:Xylosidases / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*
D014990:Xylans; D002482:Cellulose; D014995:Xylosidases; D043325:Xylan Endo-1,3-beta-Xylosidase