title: Mechanistic insight into the substrate specificity of 1,2-β-oligoglucan phosphorylase from Lachnoclostridium phytofermentans.
abstract: Glycoside phosphorylases catalyze the phosphorolysis of oligosaccharides into sugar phosphates. Recently, we found a novel phosphorylase acting on β-1,2-glucooligosaccharides with degrees of polymerization of 3 or more (1,2-β-oligoglucan phosphorylase, SOGP) in glycoside hydrolase family (GH) 94. Here, we characterized SOGP from Lachnoclostridium phytofermentans (LpSOGP) and determined its crystal structure. LpSOGP is a monomeric enzyme that contains a unique β-sandwich domain (Ndom1) at its N-terminus. Unlike the dimeric GH94 enzymes possessing catalytic pockets at their dimer interface, LpSOGP has a catalytic pocket between Ndom1 and the catalytic domain. In the complex structure of LpSOGP with sophorose, sophorose binds at subsites +1 to +2. Notably, the Glc moiety at subsite +1 is flipped compared with the corresponding ligands in other GH94 enzymes. This inversion suggests the great distortion of the glycosidic bond between subsites -1 and +1, which is likely unfavorable for substrate binding. Compensation for this disadvantage at subsite +2 can be accounted for by the small distortion of the glycosidic bond in the sophorose molecule. Therefore, the binding mode at subsites +1 and +2 defines the substrate specificity of LpSOGP, which provides mechanistic insights into the substrate specificity of a phosphorylase acting on β-1,2-glucooligosaccharides.
authors.affiliation: Department of Applied Biological Science, Faculty of Science and Technology, Tokyo University of Science, Chiba, Japan.
authors.forename: Masahiro
authors.initials: M
authors.lastname: Nakajima
authors.affiliation: Department of Applied Biological Science, Faculty of Science and Technology, Tokyo University of Science, Chiba, Japan.
authors.forename: Nobukiyo
authors.initials: N
authors.lastname: Tanaka
authors.affiliation: Department of Applied Biological Science, Faculty of Science and Technology, Tokyo University of Science, Chiba, Japan.|Department of Applied Life Sciences, Niigata University of Pharmacy and Applied Life Sciences, Niigata, Japan.
authors.forename: Nayuta
authors.initials: N
authors.lastname: Furukawa
authors.affiliation: Graduate School of Science &Technology, Niigata University, Niigata, Japan.
authors.forename: Takanori
authors.initials: T
authors.lastname: Nihira
authors.affiliation: Department of Applied Biological Science, Faculty of Science and Technology, Tokyo University of Science, Chiba, Japan.
authors.forename: Yuki
authors.initials: Y
authors.lastname: Kodutsumi
authors.affiliation: Graduate School of Science &Technology, Niigata University, Niigata, Japan.
authors.forename: Yuta
authors.initials: Y
authors.lastname: Takahashi
authors.affiliation: Graduate School of Science &Technology, Niigata University, Niigata, Japan.
authors.forename: Naohisa
authors.initials: N
authors.lastname: Sugimoto
authors.affiliation: Department of Chemistry, Tokyo Institute of Technology, Tokyo, Japan.
authors.forename: Akimasa
authors.initials: A
authors.lastname: Miyanaga
authors.affiliation: Department of Biotechnology, The University of Tokyo, Tokyo, Japan.
authors.forename: Shinya
authors.initials: S
authors.lastname: Fushinobu
authors.affiliation: Department of Applied Biological Science, Faculty of Science and Technology, Tokyo University of Science, Chiba, Japan.
authors.forename: Hayao
authors.initials: H
authors.lastname: Taguchi
authors.affiliation: Graduate School of Science &Technology, Niigata University, Niigata, Japan.
authors.forename: Hiroyuki
authors.initials: H
authors.lastname: Nakai
mesh_terms: D000595:Amino Acid Sequence; D001665:Binding Sites; D001669:Biochemical Phenomena; D020134:Catalytic Domain; D003013:Clostridium / Q000201:enzymology*; D006026:Glycoside Hydrolases / Q000737:chemistry* / Q000378:metabolism*; D006860:Hydrogen Bonding; D007700:Kinetics; D008024:Ligands; D055595:Mechanical Phenomena; D008958:Models, Molecular; D011485:Protein Binding; D011487:Protein Conformation; D000072417:Protein Domains; D054730:Protein Interaction Domains and Motifs; D013329:Structure-Activity Relationship; D013379:Substrate Specificity
chemical_list: D008024:Ligands; D006026:Glycoside Hydrolases