title: Characterization of an exo-β-1,3-D: -galactanase from Sphingomonas sp. 24T and its application to structural analysis of larch wood arabinogalactan.
abstract: A type II arabinogalactan-degrading enzyme, termed Exo-1,3-Gal, was purified to homogeneity from the culture filtrate of Sphingomonas sp. 24T. It has an apparent molecular mass of 48 kDa by SDS-PAGE. Exo-1,3-Gal was stable from pH 3 to 10 and at temperatures up to 40 °C. The optimum pH and temperature for enzyme activity were pH 6 to 7 and 50 °C, respectively. Galactose was released from β-1,3-D: -galactan and β-1,3-D: -galactooligosaccharides by the action of Exo-1,3-Gal, indicating that the enzyme was an exo-β-1,3-D: -galactanase. Analysis of the reaction products of β-1,3-galactotriose by high-performance anion-exchange chromatography revealed that the enzyme hydrolyzed the substrate in a non-processive mode. Exo-1,3-Gal bypassed the branching points of β-1,3-galactan backbones in larch wood arabinogalactan (LWAG) to produce mainly galactose, β-1,6-galactobiose, and unidentified oligosaccharides 1 and 2 with the molar ratios of 7:19:62:12. Oligosaccharides 1 and 2 were enzymatically determined to be β-1,6-galactotriose and β-1,6-galactotriose substituted with a single arabinofuranose residue, respectively. The ratio of side chains enzymatically released from LWAG was in good agreement with the postulated structure of the polysaccharide previously determined by chemical methods.
authors: Division of Applied Life Sciences, Graduate School of Life and Environmental Sciences, Osaka Prefecture University, Sakai, Osaka, Japan. sakamoto@biochem.osakafu-u.ac.jp
authors: Tatsuji
authors: T
authors: Sakamoto
authors: Hiromasa
authors: H
authors: Tanaka
authors: Yuichi
authors: Y
authors: Nishimura
authors: Megumi
authors: M
authors: Ishimaru
authors: Naoya
authors: N
authors: Kasai
mesh_terms: D001426:Bacterial Proteins / Q000737:chemistry* / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism; D004795:Enzyme Stability; D005685:Galactans / Q000737:chemistry* / Q000378:metabolism; D006026:Glycoside Hydrolases / Q000737:chemistry* / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism; D007700:Kinetics; D028221:Larix / Q000737:chemistry*; D008969:Molecular Sequence Data; D020578:Sphingomonas / Q000737:chemistry / Q000201:enzymology* / Q000235:genetics; D013379:Substrate Specificity; D014934:Wood / Q000737:chemistry*
chemical_list: D001426:Bacterial Proteins; D005685:Galactans; D006026:Glycoside Hydrolases; C063901:exo-beta-(1--3)-D-galactanase; C005653:arabinogalactan