Gene cloning, expression, and characterization of a family 51 alpha-L-arabinofuranosidase from Streptomyces sp. S9.
An alpha-L-arabinofuranosidase gene, abf51S9, was cloned from Streptomyces sp. S9 and successfully expressed in Escherichia coli BL21 (DE3). The full-length gene consisted of 1,506 bp and encoded 501 amino acids with a calculated mass of 55.2 kDa. The deduced amino acid sequence was highly homologous with the alpha-L-arabinofuranosidases belonging to family 51 of the glycoside hydrolases. The recombinant protein was purified to electrophoretic homogeneity by Ni-NTA affinity chromatography and subsequently characterized. The optimal pH and temperature for the recombinant enzyme were 6.0 and 60 approximately 65 degrees C, respectively. The enzyme showed a broad pH range of stability, retaining over 75% of the maximum activity at pH 5.0 to 11.0. The specific activity, K (m), and V (max) with p-nitrophenyl-alpha-L-arabinofuranoside as substrate were 60.0 U mg(-1), 1.45 mM, and 221 mumol min(-1) mg(-1), respectively. Abf51S9 showed a mild but significant synergistic effect in combination with xylanase on the degradation of oat-spelt xylan and soluble wheat arabinoxylan substrates with a 1.19- and 1.21-fold increase in the amount of reducing sugar released, respectively. These favorable properties make Abf51S9 a good candidate in various industrial applications.
Key Laboratory for Feed Biotechnology of Ministry of Agriculture, Feed Research Institute, Chinese Academy of Agricultural Sciences, No. 12 Zhongguancun South Street, Beijing 100081, People's Republic of China.
Pengjun
P
Shi
Ning
N
Li
Peilong
P
Yang
Yaru
Y
Wang
Huiying
H
Luo
Yingguo
Y
Bai
Bin
B
Yao
D000595:Amino Acid Sequence; D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism; D002846:Chromatography, Affinity; D003001:Cloning, Molecular; D006026:Glycoside Hydrolases / Q000235:genetics / Q000378:metabolism*; D006863:Hydrogen-Ion Concentration; D008969:Molecular Sequence Data; D017386:Sequence Homology, Amino Acid; D013302:Streptomyces / Q000201:enzymology*; D014990:Xylans / Q000378:metabolism
D001426:Bacterial Proteins; D014990:Xylans; D006026:Glycoside Hydrolases; C019713:alpha-N-arabinofuranosidase