title: Structural and biochemical analyses of glycoside hydrolase family 26 β-mannanase from a symbiotic protist of the termite Reticulitermes speratus.
abstract: Termites and their symbiotic protists have established a prominent dual lignocellulolytic system, which can be applied to the biorefinery process. One of the major components of lignocellulose from conifers is glucomannan, which comprises a heterogeneous combination of β-1,4-linked mannose and glucose. Mannanases are known to hydrolyze the internal linkage of the glucomannan backbone, but the specific mechanism by which they recognize and accommodate heteropolysaccharides is currently unclear. Here, we report biochemical and structural analyses of glycoside hydrolase family 26 mannanase C (RsMan26C) from a symbiotic protist of the termite Reticulitermes speratus. RsMan26C was characterized based on its catalytic efficiency toward glucomannan, compared with pure mannan. The crystal structure of RsMan26C complexed with gluco-manno-oligosaccharide(s) explained its specificities for glucose and mannose at subsites -5 and -2, respectively, in addition to accommodation of both glucose and mannose at subsites -3 and -4. RsMan26C has a long open cleft with a hydrophobic platform of Trp(94) at subsite -5, facilitating enzyme binding to polysaccharides. Notably, a unique oxidized Met(85) specifically interacts with the equatorial O-2 of glucose at subsite -3. Our results collectively indicate that specific recognition and accommodation of glucose at the distal negative subsites confers efficient degradation of the heteropolysaccharide by mannanase.
authors.affiliation: Department of Biotechnology, Graduate School of Agricultural and Life Sciences, The University of Tokyo, Tokyo 113-8657, Japan.
authors.forename: Hikaru
authors.initials: H
authors.lastname: Tsukagoshi
authors.affiliation: Department of Biomaterials Sciences, Graduate School of Agricultural and Life Sciences, The University of Tokyo, Tokyo 113-8657, Japan.
authors.forename: Akihiko
authors.initials: A
authors.lastname: Nakamura
authors.affiliation: Department of Biomaterials Sciences, Graduate School of Agricultural and Life Sciences, The University of Tokyo, Tokyo 113-8657, Japan.
authors.forename: Takuya
authors.initials: T
authors.lastname: Ishida
authors.affiliation: Department of Biotechnology, Graduate School of Agricultural and Life Sciences, The University of Tokyo, Tokyo 113-8657, Japan.
authors.forename: Kouki K
authors.initials: KK
authors.lastname: Touhara
authors.affiliation: Biomass Engineering Program Cooperation Division, RIKEN, Yokohama, Kanagawa 230-0045, Japan.
authors.forename: Masato
authors.initials: M
authors.lastname: Otagiri
authors.affiliation: Laboratory of Environmental Molecular Biology, Graduate School of Yokohama City University, Yokohama, Kanagawa 230-0045, Japan; Molecular and Informative Life Science Unit, RIKEN Advanced Science Institute, Yokohama, Kanagawa 230-0045, Japan.
authors.forename: Shigeharu
authors.initials: S
authors.lastname: Moriya
authors.affiliation: Department of Biomaterials Sciences, Graduate School of Agricultural and Life Sciences, The University of Tokyo, Tokyo 113-8657, Japan.
authors.forename: Masahiro
authors.initials: M
authors.lastname: Samejima
authors.affiliation: Department of Biomaterials Sciences, Graduate School of Agricultural and Life Sciences, The University of Tokyo, Tokyo 113-8657, Japan.
authors.forename: Kiyohiko
authors.initials: K
authors.lastname: Igarashi
authors.affiliation: Department of Biotechnology, Graduate School of Agricultural and Life Sciences, The University of Tokyo, Tokyo 113-8657, Japan.
authors.forename: Shinya
authors.initials: S
authors.lastname: Fushinobu
authors.affiliation: Department of Biotechnology, Graduate School of Agricultural and Life Sciences, The University of Tokyo, Tokyo 113-8657, Japan.
authors.forename: Katsuhiko
authors.initials: K
authors.lastname: Kitamoto
authors.affiliation: Department of Biotechnology, Graduate School of Agricultural and Life Sciences, The University of Tokyo, Tokyo 113-8657, Japan. Electronic address: arioka@mail.ecc.u-tokyo.ac.jp.
authors.forename: Manabu
authors.initials: M
authors.lastname: Arioka
mesh_terms: D000818:Animals; D018533:Biomass; D002384:Catalysis; D020134:Catalytic Domain; D002855:Chromatography, Thin Layer; D056890:Eukaryota / Q000201:enzymology; D005947:Glucose / Q000378:metabolism; D006868:Hydrolysis; D007422:Intestines / Q000382:microbiology; D020049:Isoptera / Q000382:microbiology*; D008031:Lignin / Q000378:metabolism; D008351:Mannans / Q000378:metabolism*; D008358:Mannose / Q000378:metabolism; D008361:Mannosidases / Q000378:metabolism*; D011134:Polysaccharides / Q000378:metabolism; D011487:Protein Conformation; D013379:Substrate Specificity; D013559:Symbiosis; D044902:beta-Mannosidase / Q000378:metabolism*
chemical_list: D008351:Mannans; D011134:Polysaccharides; C036909:lignocellulose; C022901:(1-6)-alpha-glucomannan; D008031:Lignin; D008361:Mannosidases; D044902:beta-Mannosidase; D005947:Glucose; D008358:Mannose
keywords: Crystal Structure; Glycoside Hydrolases; Mannanase; Plant Cell Wall; Polysaccharide; Symbiosis; Symbiotic Protist; Termite