title: Purification, characterization, and substrate specificity of a glucoamylase with steroidal saponin-rhamnosidase activity from Curvularia lunata.
abstract: It has been previously reported that a glucoamylase from Curvularia lunata is able to hydrolyze the terminal 1,2-linked rhamnosyl residues of sugar chains at C-3 position of steroidal saponins. In this work, the enzyme was isolated and identified after isolation and purification by column chromatography including gel filtration and ion-exchange chromatography. Analysis of protein fragments by MALDI-TOF/TOF proteomics Analyzer indicated the enzyme to be 1,4-alpha-D-glucan glucohydrolase EC 3.2.1.3, GA and had considerable homology with the glucoamylase from Aspergillus oryzae. We first found that the glucoamylase was produced from C. lunata and was able to hydrolyze the terminal rhamnosyl of steroidal saponins. The enzyme had the general character of glucoamylase, which hydrolyze starch. It had a molecular mass of 66 kDa and was optimally active at 50 degrees C, pH 4, and specific activity of 12.34 U mg of total protein(-1) under the conditions, using diosgenin-3-O-alpha-L-rhamnopyranosyl(1-->4)-[alpha-L-rhamnopyranosyl (1-->2)]-beta-D-glucopyranoside (compound II) as the substrate. Furthermore, four kinds of commercial glucoamylases from Aspergillus niger were investigated in this work, and they had the similar activity in hydrolyzing terminal rhamnosyl residues of steroidal saponin.
authors.affiliation: Beijing Institute of Radiation Medicine, Beijing 100850, China. istas1101@163.com
authors.forename: Bing
authors.initials: B
authors.lastname: Feng
authors.forename: Wei
authors.initials: W
authors.lastname: Hu
authors.forename: Bai-ping
authors.initials: BP
authors.lastname: Ma
authors.forename: Yong-ze
authors.initials: YZ
authors.lastname: Wang
authors.forename: Hong-ze
authors.initials: HZ
authors.lastname: Huang
authors.forename: Sheng-qi
authors.initials: SQ
authors.lastname: Wang
authors.forename: Xiao-hong
authors.initials: XH
authors.lastname: Qian
mesh_terms: D000595:Amino Acid Sequence; D004795:Enzyme Stability; D005087:Glucan 1,4-alpha-Glucosidase / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D006026:Glycoside Hydrolases / Q000378:metabolism*; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D003904:Mitosporic Fungi / Q000201:enzymology*; D008969:Molecular Sequence Data; D015394:Molecular Structure; D012503:Saponins / Q000378:metabolism*; D013379:Substrate Specificity; D013696:Temperature