title: A Fungal α-Galactosidase from Tricholoma matsutake with Broad Substrate Specificity and Good Hydrolytic Activity on Raffinose Family Oligosaccharides.
abstract: An acidic α-galactosidase designated as TMG was purified from the fruiting bodies The purification protocol entailed ion exchange chromatography on Q-Sepharose and of Tricholoma matsutake with 136-fold purification and a specific activity of 909 units/mg. Mono-Q and fast protein liquid chromatography on Superdex 75. TMG is a monomeric protein exhibiting a molecular mass of 47 kDa in SDS-PAGE and gel filtration. The purified enzyme was identified by LC-MS/MS and three inner amino acid sequences were obtained. The optimum pH and temperature for TMG with pNPGal as substrate were pH 4.5 and 55 °C, respectively. The α-galactosidase activity was strongly inhibited by K+, Ca2+, Cd2+, Hg2+, Ag+ and Zn2+ ions. The enzyme activity was inhibited by the chemical modification agent N-bromosuccinimide (NBS), indicating the importance of tryptophan residue(s) at or near the active site. Besides hydrolyzing pNPGal, TMG also efficaciously catalyzed the degradation of natural substrates such as stachyose, raffinose, and melibiose. Thus TMG can be exploited commercially for improving the nutritional value of soy milk by degradation of indigestible oligosaccharides.
authors.affiliation: State Key Laboratory for Agrobiotechnology, Department of Microbiology, China Agricultural University, Beijing 100193, China. gengxueran2007@163.com.
authors.forename: Xueran
authors.initials: X
authors.lastname: Geng
authors.affiliation: Institute of Biotechnology and Germplasmic Resource, Yunnan Academy of Agricultural Science, Kunming 650223, China. tiangt@aliyun.com.
authors.forename: Guoting
authors.initials: G
authors.lastname: Tian
authors.affiliation: Institute of Biotechnology and Germplasmic Resource, Yunnan Academy of Agricultural Science, Kunming 650223, China. yaasmushroom@aliyun.com.
authors.forename: Yongchang
authors.initials: Y
authors.lastname: Zhao
authors.affiliation: College of Food Science and Technology, Nanjing Agricultural University, Weigang, Nanjing 210095, China. zhlychen@njau.edu.cn.
authors.forename: Liyan
authors.initials: L
authors.lastname: Zhao
authors.affiliation: State Key Laboratory for Agrobiotechnology, Department of Microbiology, China Agricultural University, Beijing 100193, China. hxwang@cau.edu.cn.
authors.forename: Hexiang
authors.initials: H
authors.lastname: Wang
authors.affiliation: School of Biomedical Sciences, Faculty of Medicine, The Chinese University of Hong Kong, Shatin, New Territories, Hong Kong, China. b021770@mailserv.cuhk.edu.hk.
authors.forename: Tzi Bun
authors.initials: TB
authors.lastname: Ng
mesh_terms: D005656:Fungal Proteins / Q000737:chemistry / Q000302:isolation & purification; D009844:Oligosaccharides / Q000737:chemistry*; D011887:Raffinose / Q000737:chemistry*; D013379:Substrate Specificity; D055439:Tricholoma / Q000201:enzymology*; D000519:alpha-Galactosidase / Q000737:chemistry / Q000302:isolation & purification
chemical_list: D005656:Fungal Proteins; D009844:Oligosaccharides; D000519:alpha-Galactosidase; D011887:Raffinose
keywords: Tricholoma matsutake; characterization; mushroom; álpha-galactosidase