title: A novel xylan degrading β-D-xylosidase: purification and biochemical characterization.
abstract: Aspergillus ochraceus, a thermotolerant fungus isolated in Brazil from decomposing materials, produced an extracellular β-xylosidase that was purified using DEAE-cellulose ion exchange chromatography, Sephadex G-100 and Biogel P-60 gel filtration. β-xylosidase is a glycoprotein (39 % carbohydrate content) and has a molecular mass of 137 kDa by SDS-PAGE, with optimal temperature and pH at 70 °C and 3.0-5.5, respectively. β-xylosidase was stable in acidic pH (3.0-6.0) and 70 °C for 1 h. The enzyme was activated by 5 mM MnCl₂ (28 %) and MgCl₂ (20 %) salts. The β-xylosidase produced by A. ochraceus preferentially hydrolyzed p-nitrophenyl-β-D-xylopyranoside, exhibiting apparent K(m) and V(max) values of 0.66 mM and 39 U (mg protein)⁻¹ respectively, and to a lesser extent p-nitrophenyl-β-D-glucopyranoside. The enzyme was able to hydrolyze xylan from different sources, suggesting a novel β-D-xylosidase that degrades xylan. HPLC analysis revealed xylans of different compositions which allowed explaining the differences in specificity observed by β-xylosidase. TLC confirmed the capacity of the enzyme in hydrolyzing xylan and larger xylo-oligosaccharides, as xylopentaose.
authors: Departamento de Biologia, Faculdade de Filosofia, Ciências e Letras de Ribeirão Preto, Universidade de São Paulo, Avenida Bandeirantes 3900, Ribeirão Preto, SP 14040-901, Brazil.
authors: Michele
authors: M
authors: Michelin
authors: Simone C
authors: SC
authors: Peixoto-Nogueira
authors: Tony M
authors: TM
authors: Silva
authors: João A
authors: JA
authors: Jorge
authors: Héctor F
authors: HF
authors: Terenzi
authors: José A
authors: JA
authors: Teixeira
authors: Maria de Lourdes T M
authors: Mde L
authors: Polizeli
mesh_terms: D001235:Aspergillus ochraceus / Q000201:enzymology* / Q000302:isolation & purification; D001938:Brazil; D002712:Chlorides / Q000378:metabolism; D002850:Chromatography, Gel; D002852:Chromatography, Ion Exchange; D004783:Environmental Microbiology; D020536:Enzyme Activators / Q000378:metabolism; D004795:Enzyme Stability; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D015636:Magnesium Chloride / Q000378:metabolism; D017895:Manganese Compounds / Q000378:metabolism; D008970:Molecular Weight; D013379:Substrate Specificity; D013696:Temperature; D014990:Xylans / Q000378:metabolism*; D014995:Xylosidases / Q000737:chemistry / Q000302:isolation & purification* / Q000378:metabolism*
chemical_list: D002712:Chlorides; D020536:Enzyme Activators; D017895:Manganese Compounds; D014990:Xylans; D015636:Magnesium Chloride; D014995:Xylosidases; C026579:exo-1,4-beta-D-xylosidase; C025340:manganese chloride