Cloning, functional expression and characterization of three Phanerochaete chrysosporium endo-1,4-beta-xylanases.
Three Phanerochaete chrysosporium endo-1,4-beta-xylanase genes were cloned and expressed in Aspergillus niger. Two of these genes, xynA and xynC, encode family 10 glycoside hydrolases, while the third, xynB, codes for a family 11 glycoside hydrolase. All three xylanases possess a type I carbohydrate-binding domain connected to the catalytic domain by a linker region. The three xylanases were purified to homogeneity by weak anion or Avicell column chromatography and subsequently characterized. The XynA, XynB and XynC enzymes have molecular masses of 52, 30 and 50 kDa, respectively. Optimal activity was obtained at pH 4.5 and 70 degrees C with the family 10 xylanases and pH 4.5 and 60 degrees C with the family 11 xylanase. The measured Km when using birchwood xylan as the substrate was 3.71 +/- 0.69 mg/ml for XynA and XynC and was 9.96 +/- 1.45 mg/ml for XynB. Substrate specificity studies and the products released during the degradation of birchwood xylan suggest differences in catalytic properties between the two family 10 xylanases and the family 11 xylanase.
Centre for Structural and Functional Genomics, Department of Biology, Concordia University, 7141 Sherbrooke W, Montreal, Quebec, Canada.
Barbara
B
Decelle
Adrian
A
Tsang
Reginald K
RK
Storms
D000595:Amino Acid Sequence; D001234:Aspergillus niger; D002384:Catalysis; D002855:Chromatography, Thin Layer; D003001:Cloning, Molecular; D019295:Computational Biology; D017931:DNA Primers; D043364:Endo-1,4-beta Xylanases / Q000235:genetics* / Q000378:metabolism; D015966:Gene Expression Regulation, Fungal; D006026:Glycoside Hydrolases / Q000235:genetics* / Q000378:metabolism; D006863:Hydrogen-Ion Concentration; D005810:Multigene Family / Q000235:genetics; D020075:Phanerochaete / Q000201:enzymology / Q000235:genetics*; D011487:Protein Conformation; D016415:Sequence Alignment; D013379:Substrate Specificity / Q000235:genetics; D013696:Temperature; D014170:Transformation, Genetic
D017931:DNA Primers; D006026:Glycoside Hydrolases; D043364:Endo-1,4-beta Xylanases