title: Characterization of a protease-resistant α-galactosidase from the thermophilic fungus Rhizomucor miehei and its application in removal of raffinose family oligosaccharides.
abstract: The α-galactosidase gene, RmGal36, from Rhizomucor miehei was cloned and expressed in Escherichia coli. The gene has an open reading frame of 2256bp encoding 751 amino acid residues. RmGal36 was optimally active at pH 4.5 and 60°C, but is stable between pH 4.5 and 10.0 and at a temperature of up to 55°C for 30min retaining more than 80% of its relative activity. It displayed remarkable resistance to proteases and its activity was not inhibited by galactose concentrations of 100mM. The relative specificity of RmGal36 towards various substrates is in the order of p-nitrophenyl α-galactopyranoside>melibiose>stachyose>raffinose, with a K(m) of 0.36, 16.9, 27.6, and 47.9mM, respectively. The enzyme completely hydrolyzed raffinose and stachyose present in soybeans and kidney beans at 50°C within 60min. These features make RmGal36 useful in the food and feed industries and in processing of beet-sugar.
authors.affiliation: Department of Biotechnology, College of Food Science and Nutritional Engineering, China Agricultural University, Beijing 100083, China.
authors.forename: Priti
authors.initials: P
authors.lastname: Katrolia
authors.forename: Huiyong
authors.initials: H
authors.lastname: Jia
authors.forename: Qiaojuan
authors.initials: Q
authors.lastname: Yan
authors.forename: Shuang
authors.initials: S
authors.lastname: Song
authors.forename: Zhengqiang
authors.initials: Z
authors.lastname: Jiang
authors.forename: Haibo
authors.initials: H
authors.lastname: Xu
mesh_terms: D000595:Amino Acid Sequence; D001483:Base Sequence; D018076:DNA, Complementary; D004591:Electrophoresis, Polyacrylamide Gel; D004795:Enzyme Stability; D006358:Hot Temperature; D006863:Hydrogen-Ion Concentration; D006868:Hydrolysis; D007700:Kinetics; D008969:Molecular Sequence Data; D010447:Peptide Hydrolases / Q000378:metabolism*; D011887:Raffinose / Q000302:isolation & purification*; D020103:Rhizomucor / Q000201:enzymology*; D013379:Substrate Specificity; D000519:alpha-Galactosidase / Q000378:metabolism*
chemical_list: D018076:DNA, Complementary; D000519:alpha-Galactosidase; D010447:Peptide Hydrolases; D011887:Raffinose