Purification and characterisation of a major xylanase with cellulase and transferase activities from Fusarium oxysporum.
A major xylanase from Fusarium oxysporum was purified to homogeneity by gel filtration, affinity, and ion-exchange chromatographies. It has a molecular mass of 60.2 kDa and pI of 6.6 and was optimally active at pH 7.4 and at 50 degrees C. The enzyme was stable over the pH range 5.8-8.2 at 40 degrees C for 24 h and lost 45% of its original activity at pH 9.0 under the identical conditions. The enzyme rapidly hydrolysed xylans from oat spelts (husks) and birchwood, but the activities on carboxymethylcellulose (CMC), filter paper, and Avicel were very low. Determination of kcat/Km revealed that the enzyme hydrolysed oat spelts and birchwood xylans, 15-30 times more efficiently than CMC. In a 24 h incubation, at pH 7.0 and 9.0, the enzyme hydrolysed oat spelts and birchwood xylans by 75 and 65%, respectively. However, at pH 7.0, the enzyme released almost equal amounts of xylose and xylobiose from both xylans, whereas at pH 9.0, the concentration of xylobiose was twice as muchi as that of xylose and xylotriose. Xylanase attacked preferentially the internal glycosidic bonds of xylo- and 4-methylumbelliferyl cello-oligosaccharides [MeUmb(Glc)n]. The enzyme catalysed transglycosylation reaction with xylotriose, xylotetraose, and xylopentaose as donors and 4-methylumbelliferyl beta-D-glucoside (MeUmbGlc) as an acceptor.
Christakopoulos
P
P
Food Macromolecular Science Department, Institute of Food Research, Reading Laboratory, UK.
Kekos
D
D
Macris
B J
BJ
Claeyssens
M
M
Bhat
M K
MK
D002480:Cellulase / Q000378:metabolism; D002850:Chromatography, Gel; D002851:Chromatography, High Pressure Liquid; D004591:Electrophoresis, Polyacrylamide Gel; D004795:Enzyme Stability; D005110:Extracellular Space / Q000737:chemistry / Q000201:enzymology / Q000378:metabolism; D005670:Fusarium / Q000737:chemistry / Q000201:enzymology*; D006027:Glycosides / Q000378:metabolism; D016695:Glycosyltransferases / Q000378:metabolism; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D011134:Polysaccharides / Q000378:metabolism; D013379:Substrate Specificity; D013696:Temperature; D014468:Umbelliferones / Q000378:metabolism; D043325:Xylan Endo-1,3-beta-Xylosidase; D014990:Xylans / Q000378:metabolism; D014995:Xylosidases / Q000737:chemistry / Q000302:isolation & purification / Q000378:metabolism*
D006027:Glycosides; D011134:Polysaccharides; D014468:Umbelliferones; D014990:Xylans; D016695:Glycosyltransferases; D014995:Xylosidases; D043325:Xylan Endo-1,3-beta-Xylosidase; D002480:Cellulase