title: A Novel Glycoside Hydrolase Family 113 Endo-β-1,4-Mannanase from Alicyclobacillus sp. Strain A4 and Insight into the Substrate Recognition and Catalytic Mechanism of This Family.
abstract: Few members of glycoside hydrolase (GH) family 113 have been characterized, and information on substrate recognition by and the catalytic mechanism of this family is extremely limited. In the present study, a novel endo-β-1,4-mannanase of GH 113, Man113A, was identified in thermoacidophilic Alicyclobacillus sp. strain A4 and found to exhibit both hydrolytic and transglycosylation activities. The enzyme had a broad substrate spectrum, showed higher activities on glucomannan than on galactomannan, and released mannobiose and mannotriose as the main hydrolysis products after an extended incubation. Compared to the only functionally characterized and structure-resolved counter part Alicyclobacillus acidocaldarius ManA (AaManA) of GH 113, Man113A showed much higher catalytic efficiency on mannooligosaccharides, in the order mannohexaose ≈ mannopentaose > mannotetraose > mannotriose, and required at least four sugar units for efficient catalysis. Homology modeling, molecular docking analysis, and site-directed mutagenesis revealed the vital roles of eight residues (Trp13, Asn90, Trp96, Arg97, Tyr196, Trp274, Tyr292, and Cys143) related to substrate recognition by and catalytic mechanism of GH 113. Comparison of the binding pockets and key residues of β-mannanases of different families indicated that members of GH 113 and GH 5 have more residues serving as stacking platforms to support -4 to -1 subsites than those of GH 26 and that the residues preceding the acid/base catalyst are quite different. Taken as a whole, this study elucidates substrate recognition by and the catalytic mechanism of GH 113 β-mannanases and distinguishes them from counterparts of other families.
authors.affiliation: Key Laboratory for Feed Biotechnology of the Ministry of Agriculture, Feed Research Institute, Chinese Academy of Agricultural Sciences, Beijing, People's Republic of China.|College of Animal Science, Zhejiang University, Hangzhou, People's Republic of China.
authors.forename: Wei
authors.initials: W
authors.lastname: Xia
authors.affiliation: College of Food Science and Technology, Hebei Agricultural University, Baoding, People's Republic of China.
authors.forename: Haiqiang
authors.initials: H
authors.lastname: Lu
authors.affiliation: Key Laboratory for Feed Biotechnology of the Ministry of Agriculture, Feed Research Institute, Chinese Academy of Agricultural Sciences, Beijing, People's Republic of China.
authors.forename: Mengjuan
authors.initials: M
authors.lastname: Xia
authors.affiliation: Key Laboratory for Feed Biotechnology of the Ministry of Agriculture, Feed Research Institute, Chinese Academy of Agricultural Sciences, Beijing, People's Republic of China.
authors.forename: Ying
authors.initials: Y
authors.lastname: Cui
authors.affiliation: Key Laboratory for Feed Biotechnology of the Ministry of Agriculture, Feed Research Institute, Chinese Academy of Agricultural Sciences, Beijing, People's Republic of China.
authors.forename: Yingguo
authors.initials: Y
authors.lastname: Bai
authors.affiliation: College of Animal Science, Zhejiang University, Hangzhou, People's Republic of China.
authors.forename: Lichun
authors.initials: L
authors.lastname: Qian
authors.affiliation: Key Laboratory for Feed Biotechnology of the Ministry of Agriculture, Feed Research Institute, Chinese Academy of Agricultural Sciences, Beijing, People's Republic of China binyao@caas.cn shipengjun@caas.cn.
authors.forename: Pengjun
authors.initials: P
authors.lastname: Shi
authors.affiliation: Key Laboratory for Feed Biotechnology of the Ministry of Agriculture, Feed Research Institute, Chinese Academy of Agricultural Sciences, Beijing, People's Republic of China.
authors.forename: Huiying
authors.initials: H
authors.lastname: Luo
authors.affiliation: Key Laboratory for Feed Biotechnology of the Ministry of Agriculture, Feed Research Institute, Chinese Academy of Agricultural Sciences, Beijing, People's Republic of China binyao@caas.cn shipengjun@caas.cn.
authors.forename: Bin
authors.initials: B
authors.lastname: Yao
mesh_terms: D056450:Alicyclobacillus / Q000201:enzymology* / Q000235:genetics; D001665:Binding Sites; D002384:Catalysis; D004789:Enzyme Activation; D005690:Galactose / Q000031:analogs & derivatives; D006027:Glycosides / Q000378:metabolism; D006868:Hydrolysis; D008351:Mannans / Q000378:metabolism; D008361:Mannosidases / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D062105:Molecular Docking Simulation; D016297:Mutagenesis, Site-Directed; D009844:Oligosaccharides / Q000378:metabolism; D011994:Recombinant Proteins / Q000737:chemistry / Q000302:isolation & purification / Q000378:metabolism; D040681:Structural Homology, Protein; D013379:Substrate Specificity; D014312:Trisaccharides / Q000378:metabolism; D044902:beta-Mannosidase / Q000378:metabolism
chemical_list: D006027:Glycosides; D008351:Mannans; D009844:Oligosaccharides; D011994:Recombinant Proteins; D014312:Trisaccharides; C012990:galactomannan; C014647:mannobiose; C039922:mannotetraose; C022901:(1-6)-alpha-glucomannan; C023872:mannotriose; C048068:mannopentaose; D008361:Mannosidases; D044902:beta-Mannosidase; C041867:endo-1,4-beta-D-mannanase; D005690:Galactose