Biochemical and catalytic properties of an endoxylanase purified from the culture filtrate of Thermomyces lanuginosus ATCC 46882.
An endoxylanase (1,4-beta-D-xylan xylanohydrolase, EC 3.2.1.8) from the culture filtrates of T. lanuginosus ATCC 46882 was purified to homogeneity by DEAE-Sepharose and Bio-Gel P-30 column chromatographies. The purified endoxylanase had a specific activity of 888.8 mumol min-1 mg-1 protein and accounted for approximately 30% of the total protein secreted by this fungus. The molecular mass of native (non-denatured) and denatured endoxylanase were 26.3 and 25.7 kD as, respectively. Endoxylanase had a pI of 3.7 and was optimally active between pH 6.0-6.5 and at 75 degrees C. The enzyme showed > 50% of its original activity between pH 5.5-9.0 and at 85 degrees C. The pH and temperature stability studies revealed that this endoxylanase was almost completely stable between pH 5.0-9.0 and up to 60 degrees C for 5 h and at pH 10.0 up to 55 degrees C for 5 h. Thin-layer chromatography (TLC) analysis showed that endoxylanase released mainly xylose (Xyl) and xylobiose (Xyl2) from beechwood 4-O-methyl-D-glucuronoxylan, O-acetyl-4-O-methyl-D-glucuronoxylan and rhodymenan (a beta-(1-->3)-beta(1-->4)-xylan). Also, the enzyme released an acidic xylo-oligosaccharide from 4-O-methyl-D-glucuronoxylan, and an isomeric xylotetraose and an isomeric xylopentaose from rhodymenan. The enzyme hydrolysed [1-3H]-xylo-oligosaccharides in an endofashion, but the hydrolysis of [1-3H]-xylotriose appeared to proceed via transglycosylation. since the xylobiose was the predominant product. Endoxylanase was not active on pNPX and pNPC at 40 and 100 mM for up to 6 h, but showed some activity toward pNPX at 100 mM after 20-24 h. The results suggested that the endoxylanase from T. lanuginosus belongs to family 11.
Bennett
N A
NA
Food Macromolecular Science Department, Reading Laboratory, UK.
Ryan
J
J
Biely
P
P
Vrsanska
M
M
Kremnicky
L
L
Macris
B J
BJ
Kekos
D
D
Christakopoulos
P
P
Katapodis
P
P
Claeyssens
M
M
Nerinckx
W
W
Ntauma
P
P
Bhat
M K
MK
D004187:Disaccharides / Q000378:metabolism; D043364:Endo-1,4-beta Xylanases; D004795:Enzyme Stability; D005656:Fungal Proteins / Q000737:chemistry; D006863:Hydrogen-Ion Concentration; D007526:Isoelectric Point; D003904:Mitosporic Fungi / Q000201:enzymology*; D009844:Oligosaccharides / Q000378:metabolism; D013379:Substrate Specificity; D013696:Temperature; D014990:Xylans / Q000378:metabolism; D014994:Xylose / Q000378:metabolism; D014995:Xylosidases / Q000737:chemistry*
D004187:Disaccharides; D005656:Fungal Proteins; D009844:Oligosaccharides; D014990:Xylans; C044967:4-O-methyl glucuronoxylan; D014994:Xylose; D014995:Xylosidases; D043364:Endo-1,4-beta Xylanases; C004173:xylobiose