Thermostable xylanase from Marasmius sp.: purification and characterization.
We have screened 766 strains of fungi from the BIOTEC Culture Collection (BCC) for xylanases working in extreme pH and/or high temperature conditions, the so-called extreme xylanases. From a total number of 32 strains producing extreme xylanases, the strain BCC7928, identified by using the internal transcribed spacer (ITS) sequence of rRNA to be a Marasmius sp., was chosen for further characterization because of its high xylanolytic activity at temperature as high as 90 degrees C. The crude enzyme possessed high thermostability and pH stability. Purification of this xylanase was carried out using an anion exchanger followed by hydrophobic interaction chromatography, yielding the enzyme with >90% homogeneity. The molecular mass of the enzyme was approximately 40 kDa. The purified enzyme retained broad working pH range of 4-8 and optimal temperature of 90 degrees C. When using xylan from birchwood as substrate, it exhibits Km and Vmax values of 2.6 +/- 0.6 mg/ml and 428 +/- 26 U/mg, respectively. The enzyme rapidly hydrolysed xylans from birchwood, beechwood, and exhibited lower activity on xylan from wheatbran, or celluloses from carboxymethylcellulose and Avicel. The purified enzyme was highly stable at temperature ranges from 50 to 70 degrees C. It retained 84% of its maximal activity after incubation in standard buffer containing 1% xylan substrate at 70 degrees C for 3 h. This thermostable xylanase should therefore be useful for several industrial applications, such as agricultural, food and biofuel.
Ratanachomsri
Ukrit
U
National Center for Genetic Engineering and Biotechnology, 113 Thailand Science Park, Pathumthani 12120, Thailand.
Sriprang
Rutchadaporn
R
Sornlek
Warasirin
W
Buaban
Benchaporn
B
Champreda
Verawat
V
Tanapongpipat
Sutipa
S
Eurwilaichitr
Lily
L
D043364:Endo-1,4-beta Xylanases / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism; D004795:Enzyme Stability; D005656:Fungal Proteins / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism; D005658:Fungi / Q000201:enzymology*; D006863:Hydrogen-Ion Concentration; D013379:Substrate Specificity; D013696:Temperature; D014990:Xylans / Q000378:metabolism
D005656:Fungal Proteins; D014990:Xylans; D043364:Endo-1,4-beta Xylanases