title: Identification of a recombinant inulin fructotransferase (difructose dianhydride III forming) from Arthrobacter sp. 161MFSha2.1 with high specific activity and remarkable thermostability.
abstract: Difructose dianhydride III (DFA III) is a functional carbohydrate produced from inulin by inulin fructotransferase (IFTase, EC 4.2.2.18). In this work, an IFTase gene from Arthrobacter sp. 161MFSha2.1 was cloned and expressed in Escherachia coli. The recombinant enzyme was purified by metal affinity chromatography. It showed significant inulin hydrolysis activity, and the produced main product from inulin was determined as DFA III by nuclear magnetic resonance analysis. The molecular mass of the purified protein was calculated to be 43 and 125 kDa by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and gel filtration, respectively, suggesting the native enzyme might be a homotrimer. The recombinant enzyme showed maximal activity as 2391 units/mg at pH 6.5 and 55 °C. It displayed the highest thermostability among previously reported IFTases (DFA III forming) and was stable up to 80 °C for 4 h of incubation. The smallest substrate was determined as nystose. The conversion ratio of inulin to DFA III reached 81% when 100 g/L inulin was catalyzed by 80 nM recombinant enzyme for 20 min at pH 6.5 and 55 °C. All of these data indicated that the IFTase (DFA III forming) from Arthrobacter sp. 161MFSha2.1 had great potential for industrial DFA III production.
authors: †State Key Laboratory of Food Science and Technology, Ministry of Education, Key Laboratory of Carbohydrate Chemistry and Biotechnology, and ‡Synergetic Innovation Center of Food Safety and Nutrition, Jiangnan University, Wuxi, Jiangsu 214122, People's Republic of China.
authors: Xiao
authors: X
authors: Wang
authors: †State Key Laboratory of Food Science and Technology, Ministry of Education, Key Laboratory of Carbohydrate Chemistry and Biotechnology, and ‡Synergetic Innovation Center of Food Safety and Nutrition, Jiangnan University, Wuxi, Jiangsu 214122, People's Republic of China.
authors: Shuhuai
authors: S
authors: Yu
authors: †State Key Laboratory of Food Science and Technology, Ministry of Education, Key Laboratory of Carbohydrate Chemistry and Biotechnology, and ‡Synergetic Innovation Center of Food Safety and Nutrition, Jiangnan University, Wuxi, Jiangsu 214122, People's Republic of China.
authors: Tao
authors: T
authors: Zhang
authors: †State Key Laboratory of Food Science and Technology, Ministry of Education, Key Laboratory of Carbohydrate Chemistry and Biotechnology, and ‡Synergetic Innovation Center of Food Safety and Nutrition, Jiangnan University, Wuxi, Jiangsu 214122, People's Republic of China.
authors: Bo
authors: B
authors: Jiang
authors: †State Key Laboratory of Food Science and Technology, Ministry of Education, Key Laboratory of Carbohydrate Chemistry and Biotechnology, and ‡Synergetic Innovation Center of Food Safety and Nutrition, Jiangnan University, Wuxi, Jiangsu 214122, People's Republic of China.
authors: Wanmeng
authors: W
authors: Mu
mesh_terms: D001173:Arthrobacter / Q000201:enzymology* / Q000235:genetics; D003001:Cloning, Molecular; D004187:Disaccharides / Q000096:biosynthesis*; D004795:Enzyme Stability; D004926:Escherichia coli / Q000378:metabolism; D005632:Fructose; D015870:Gene Expression; D006602:Hexosyltransferases / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D006863:Hydrogen-Ion Concentration; D007444:Inulin / Q000378:metabolism*; D015394:Molecular Structure; D008970:Molecular Weight; D011994:Recombinant Proteins / Q000378:metabolism*; D013696:Temperature
chemical_list: D004187:Disaccharides; D011994:Recombinant Proteins; D005632:Fructose; C087758:difructose anhydride III; D007444:Inulin; D006602:Hexosyltransferases
keywords: Arthrobacter sp.; difructose dianhydride III; inulin; inulin fructotransferase; thermostability