title: Purification, characterization and synergism in autolysis of a group of 1,3-β-glucan hydrolases from the pilei of Coprinopsis cinerea fruiting bodies.
abstract: Using a combined chromatography method, we simultaneously purified three protein fractions (II-2, II-3 and II-4) with 1,3-β-glucanase activity from extraction of pilei of Coprinopsis cinerea fruiting bodies. MALDI-TOF/TOF amino acid sequencing showed that these three fractions matched a putative exo-1,3-β-glucanase, a putative glucan 1,3-β-glucosidase and a putative glycosyl hydrolase family 16 protein annotated in the C. cinerea genome, respectively; however, they were characterized as a 1,3-β-glucosidase, an exo-1,3-β-glucanase and an endo-1,3-β-glucanase, respectively, by analysis of their substrate specificities and modes of action. This study explored how these three 1,3-β-glucoside hydrolases synergistically acted on laminarin: the endo-1,3-β-glucanase hydrolysed internal glycosidic bonds of laminarin to generate 1,3-β-oligosaccharides of various lengths, the exo-1,3-β-glucanase cleaved the longer-chain laminarioligosaccharides into short-chain disaccharides, laminaribiose and gentiobiose, and the 1,3-β-glucosidase further hydrolysed laminaribiose to glucose. The remaining gentiobiose must be hydrolysed by other 1,6-β-glucosidases. Therefore, the endo-1,3-β-glucanase, exo-1,3-β-glucanase and 1,3-β-glucosidase may act synergistically to completely degrade the 1,3-β-glucan backbone of the C. cinerea cell wall during fruiting body autolysis. These three 1,3-β-glucoside hydrolases share a similar optimum pH and optimum temperature, supporting the speculation that these enzymes work together under the same conditions to degrade 1,3-β-glucan in the C. cinerea cell wall during fruiting body autolysis.
authors.affiliation: Jiangsu Key Laboratory for Microbes and Microbial Functional Genomics, Jiangsu Engineering and Technology Research Center for Industrialization of Microbial Resources, College of Life Science, Nanjing Normal University, Nanjing 210023, PR China.
authors.forename: Yajun
authors.initials: Y
authors.lastname: Zhou
authors.affiliation: Jiangsu Key Laboratory for Microbes and Microbial Functional Genomics, Jiangsu Engineering and Technology Research Center for Industrialization of Microbial Resources, College of Life Science, Nanjing Normal University, Nanjing 210023, PR China.
authors.forename: Wenming
authors.initials: W
authors.lastname: Zhang
authors.affiliation: Jiangsu Key Laboratory for Microbes and Microbial Functional Genomics, Jiangsu Engineering and Technology Research Center for Industrialization of Microbial Resources, College of Life Science, Nanjing Normal University, Nanjing 210023, PR China.
authors.forename: Zhonghua
authors.initials: Z
authors.lastname: Liu
authors.affiliation: Jiangsu Key Laboratory for Microbes and Microbial Functional Genomics, Jiangsu Engineering and Technology Research Center for Industrialization of Microbial Resources, College of Life Science, Nanjing Normal University, Nanjing 210023, PR China.
authors.forename: Jun
authors.initials: J
authors.lastname: Wang
authors.affiliation: Jiangsu Key Laboratory for Microbes and Microbial Functional Genomics, Jiangsu Engineering and Technology Research Center for Industrialization of Microbial Resources, College of Life Science, Nanjing Normal University, Nanjing 210023, PR China.
authors.forename: Sheng
authors.initials: S
authors.lastname: Yuan
mesh_terms: D000363:Agaricales / Q000201:enzymology*; D002845:Chromatography; D004795:Enzyme Stability; D048690:Fruiting Bodies, Fungal / Q000201:enzymology; D043326:Glucan 1,3-beta-Glucosidase / Q000737:chemistry / Q000302:isolation & purification* / Q000378:metabolism*; D005936:Glucans / Q000378:metabolism*; D006863:Hydrogen-Ion Concentration; D006868:Hydrolysis; D008970:Molecular Weight; D020539:Sequence Analysis, Protein; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization; D013379:Substrate Specificity; D013696:Temperature
chemical_list: D005936:Glucans; C008247:laminaran; D043326:Glucan 1,3-beta-Glucosidase