Biochemical and catalytic properties of an endoxylanase purified from the culture filtrate of Sporotrichum thermophile.
An endo-beta-1,4-xylanase (1,4-beta-D-xylan xylanoxydrolase, EC 3.2.1.8) present in culture filtrates of Sporotrichum thermophile ATCC 34628 was purified to homogeneity by Q-Sepharose and Sephacryl S-200 column chromatographies. The enzyme has a molecular mass of 25,000 Da, an isoelectric point of 6.7, and is optimally active at pH 5 and at 70 degrees C. Thin-layer chromatography (TLC) analysis showed that endo-xylanase liberates mainly xylose (Xyl) and xylobiose (Xyl2) from beechwood 4-O-methyl-D-glucuronoxylan, O-acetyl-4-O-methylglucuronoxylan and rhodymenan (a beta-(1-->4)-beta(1-->3)-xylan). Also, the enzyme releases an acidic xylo-oligosaccharide from 4-O-methyl-D-glucuronoxylan, and an isomeric xylotetraose and an isomeric xylopentaose from rhodymenan. Analysis of reaction mixtures by high performance liquid chromatography (HPLC) revealed that the enzyme cleaves preferentially the internal glycosidic bonds of xylooligosaccharides, [1-3H]-xylooligosaccharides and xylan. The enzyme also hydrolyses the 4-methylumbelliferyl glycosides of beta-xylobiose and beta-xylotriose at the second glycosidic bond adjacent to the aglycon. The endoxylanase is not active on pNPX and pNPC. The enzyme mediates a decrease in the viscosity of xylan associated with a release of only small amounts of reducing sugar. The enzyme is irreversibly inhibited by series of omega-epoxyalkyl glycosides of D-xylopyranose. The results suggest that the endoxylanase from S. thermophile has catalytic properties similar to the enzymes belonging to family 11.
Katapodis
Petros
P
Department of Chemical Engineering, National Technical University of Athens, 5 Iroon Polytechniou Ave, Zografou Campus, 15700 Athens, Greece.
Vrsanská
Maria
M
Kekos
Dimitris
D
Nerinckx
Wim
W
Biely
Peter
P
Claeyssens
Marc
M
Macris
Basil J
BJ
Christakopoulos
Paul
P
D002384:Catalysis; D020134:Catalytic Domain / Q000502:physiology; D002850:Chromatography, Gel; D002851:Chromatography, High Pressure Liquid; D002852:Chromatography, Ion Exchange; D002855:Chromatography, Thin Layer; D017077:Culture Media, Conditioned / Q000737:chemistry; D004591:Electrophoresis, Polyacrylamide Gel; D043364:Endo-1,4-beta Xylanases / Q000037:antagonists & inhibitors / Q000737:chemistry* / Q000302:isolation & purification; D004791:Enzyme Inhibitors / Q000494:pharmacology; D005960:Glucosides / Q000378:metabolism; D006027:Glycosides / Q000378:metabolism; D006863:Hydrogen-Ion Concentration; D006923:Hymecromone / Q000031:analogs & derivatives* / Q000378:metabolism; D007526:Isoelectric Point; D007700:Kinetics; D009682:Magnetic Resonance Spectroscopy; D008970:Molecular Weight; D009844:Oligosaccharides / Q000032:analysis / Q000378:metabolism; D013175:Sporothrix / Q000737:chemistry / Q000201:enzymology*; D013379:Substrate Specificity; D013696:Temperature; D014783:Viscosity; D014990:Xylans / Q000378:metabolism
D017077:Culture Media, Conditioned; D004791:Enzyme Inhibitors; D005960:Glucosides; D006027:Glycosides; C456725:O-acetyl-(4-O-methylglucurono)xylan; D009844:Oligosaccharides; D014990:Xylans; C091993:xylosyl-1-4-xylosyl-1-(4-methylumbelliferone); C008646:4-nitrophenyl beta-D-xyloside; C034170:4-nitrophenyl beta-cellobioside; C038910:glucuronoxylan; D006923:Hymecromone; C044967:4-O-methyl glucuronoxylan; D043364:Endo-1,4-beta Xylanases