Expression and characterization of a xylosidase (Bxyl) from Bacillus halodurans C-125.
A xylosidase gene, labeled as BH1068 in genome of Bacillus halodurans C-125, was successfully cloned and overexpressed in Escherichia coli JM109. The purified enzyme was thoroughly characterized and its xylosidase function was unambiguously confirmed. It has maximum activities in neutral condition and is stable over a wide range of pH (4.5-9.0). The enzyme has a broad temperature optimal (35 degrees C-45 degrees C) and is quite stable at temperature up to 45 degrees C. The unique pH and temperature profiles of the enzyme should allow a wide range of xylanolytic operational conditions. With high specific activity of 174 mU/mg protein for its artificial substrate (p-nitrophenyl-beta-xylose) and low xylose inhibition (inhibitor constant Ki = 300 mmol/L), this enzyme is among the most active and high tolerant bacterial xylosidase to xylose inhibition. Its high synergy with commercial xylanase has been demonstrated with beechwood xylan hydrolysis, achieving a hydrolysis yield of 40%. Its neutral pH optimal and high tolerance to product inhibition complements well with its fungal counterparts that are only optimal at acidic pH and susceptible to xylose inhibition. In conclusion, this enzyme has high potential in the saccharification of xylan and xylan-containing polysaccharides.
Liang
Yanli
Y
College of Agronomy and Biotechnology, Yunnan Agricultural University, Kunming 650201, China.
Li
Xingyu
X
Shin
Hyundong
H
Chen
Rachel R
RR
Mao
Zichao
Z
D000595:Amino Acid Sequence; D001407:Bacillus / Q000145:classification / Q000201:enzymology / Q000235:genetics*; D003001:Cloning, Molecular; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism; D006868:Hydrolysis; D008969:Molecular Sequence Data; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism; D013379:Substrate Specificity; D014994:Xylose / Q000378:metabolism; D014995:Xylosidases / Q000235:genetics / Q000378:metabolism*
D011994:Recombinant Proteins; D014994:Xylose; D014995:Xylosidases