Cloning, purification, and characterization of a thermostable alpha-L-arabinofuranosidase from Anoxybacillus kestanbolensis AC26Sari |
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Authors: | Canakci Sabriye Kacagan Murat Inan Kadriye Belduz Ali Osman Saha Badal C |
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Affiliation: | (1) Department of Biology, Faculty of Arts and Sciences, Karadeniz Technical University, 61080 Trabzon, Turkey;(2) Fermentation Biotechnology Research Unit, National Center for Agricultural Utilization Research, USDA-ARS, 1815 N. University St., Peoria, IL 61604, USA |
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Abstract: | The gene, AbfAC26Sari, encoding an α-l-arabinofuranosidase from Anoxybacillus kestanbolensis AC26Sari, was isolated, cloned, sequenced, and characterizated. On the basis of amino acid sequence similarities, this 57-kDa enzyme could be assigned to family 51 of the glycosyl hydrolase classification system. Characterization of the purified recombinant α-l-arabinofuranosidase produced in Escherichia coli BL21 revealed that it is active at a broad pH range (pH 4.5 to 9.0) and at a broad temperature range (45–85°C) and it has an optimum pH of 5.5 and an optimum temperature of 65°C. Kinetic experiment at 65°C with p-nitrophenyl α-l-arabinofuranoside as a substrate gave a V max and K m values of 1,019 U/mg and 0.139 mM, respectively. The enzyme had no apparent requirement of metal ions for activity, and its activity was strongly inhibited by 1 mM Cu2+ and Hg2+. The recombinant arabinofuranosidase released l-arabinose from arabinan, arabinoxylan, oat spelt xylan, arabinobiose, arabinotriose, arabinotetraose, and arabinopentaose. Endoarabinanase activity was not detected. These findings suggest that AbfAC26Sari is an exo-acting enzyme. |
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Keywords: | α - font-variant:small-caps" >l-Arabinofuranosidase Anoxybacillus kestanbolensis Arabinan Arabinoxylan Arabinooligosaccharides |
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