title: Isomaltooligosaccharide-binding structure ofPaenibacillussp. 598K cycloisomaltooligosaccharide glucanotransferase.
abstract: Paenibacillussp. 598K cycloisomaltooligosaccharide glucanotransferase (CITase), a member of glycoside hydrolase family 66 (GH66), catalyses the intramolecular transglucosylation of dextran to produce CIs with seven or more degrees of polymerization. To clarify the cyclization reaction and product specificity of the enzyme, we determined the crystal structure of PsCITase. The core structure of PsCITase consists of four structural domains: a catalytic (β/α)8-domain and three β-domains. A family 35 carbohydrate-binding module (first CBM35 region ofPaenibacillussp. 598K CITase, (PsCBM35-1)) is inserted into and protrudes from the catalytic domain. The ligand complex structure of PsCITase prepared by soaking the crystal with cycloisomaltoheptaose yielded bound sugars at three sites: in the catalytic cleft, at the joint of the PsCBM35-1 domain and at the loop region of PsCBM35-1. In the catalytic site, soaked cycloisomaltoheptaose was observed as a linear isomaltoheptaose, presumably a hydrolysed product from cycloisomaltoheptaose by the enzyme and occupied subsites -7 to -1. Beyond subsite -7, three glucose moieties of another isomaltooiligosaccharide were observed, and these positions are considered to be distal subsites -13 to -11. The third binding site is the canonical sugar-binding site at the loop region of PsCBM35-1, where the soaked cycloisomaltoheptaose is bound. The structure indicated that the concave surface between the catalytic domain and PsCBM35-1 plays a guiding route for the long-chained substrate at the cyclization reaction.
authors.affiliation: Advanced Analysis Center, National Agriculture and Food Research Organization (NARO), 2-1-2 Kannondai, Tsukuba, Ibaraki 305-8602, Japan zui@affrc.go.jp funane@affrc.go.jp.|Biomolecular Research Unit, National Institute of Agrobiological Sciences, 2-1-2 Kannondai, Tsukuba 305-8602, Japan.
authors.forename: Zui
authors.initials: Z
authors.lastname: Fujimoto
authors.affiliation: Advanced Analysis Center, National Agriculture and Food Research Organization (NARO), 2-1-2 Kannondai, Tsukuba, Ibaraki 305-8602, Japan.|Biomolecular Research Unit, National Institute of Agrobiological Sciences, 2-1-2 Kannondai, Tsukuba 305-8602, Japan.
authors.forename: Naomi
authors.initials: N
authors.lastname: Kishine
authors.affiliation: Biomolecular Research Unit, National Institute of Agrobiological Sciences, 2-1-2 Kannondai, Tsukuba 305-8602, Japan.
authors.forename: Nobuhiro
authors.initials: N
authors.lastname: Suzuki
authors.affiliation: Department of Biological Production, Faculty of Bioresource Sciences, Akita Prefectural University, Shimoshinjyo-Nakano, Akita 010-0195, Japan.|Division of Food Biotechnology, Food Research Institute, National Agriculture and Food Research Organization, 2-1-12 Kannondai, Tsukuba 305-8642, Japan.
authors.forename: Ryuichiro
authors.initials: R
authors.lastname: Suzuki
authors.affiliation: Division of Food Biotechnology, Food Research Institute, National Agriculture and Food Research Organization, 2-1-12 Kannondai, Tsukuba 305-8642, Japan.
authors.forename: Daiki
authors.initials: D
authors.lastname: Mizushima
authors.affiliation: Biomolecular Research Unit, National Institute of Agrobiological Sciences, 2-1-2 Kannondai, Tsukuba 305-8602, Japan.|Department of Planning and General Administration, Institute of Crop Science, National Agriculture and Food Research Organization, 2-1-2 Kannondai, Tsukuba 305-8602, Japan.
authors.forename: Mitsuru
authors.initials: M
authors.lastname: Momma
authors.affiliation: Division of Food Biotechnology, Food Research Institute, National Agriculture and Food Research Organization, 2-1-12 Kannondai, Tsukuba 305-8642, Japan.
authors.forename: Keitarou
authors.initials: K
authors.lastname: Kimura
authors.affiliation: Division of Food Biotechnology, Food Research Institute, National Agriculture and Food Research Organization, 2-1-12 Kannondai, Tsukuba 305-8642, Japan zui@affrc.go.jp funane@affrc.go.jp.
authors.forename: Kazumi
authors.initials: K
authors.lastname: Funane
mesh_terms: D000595:Amino Acid Sequence; D001665:Binding Sites; D020134:Catalytic Domain; D018360:Crystallography, X-Ray; D005964:Glucosyltransferases / Q000737:chemistry* / Q000378:metabolism*; D007534:Isomaltose / Q000378:metabolism*; D008958:Models, Molecular; D056507:Paenibacillus / Q000737:chemistry / Q000201:enzymology* / Q000378:metabolism; D011485:Protein Binding; D011487:Protein Conformation; D013379:Substrate Specificity
chemical_list: D007534:Isomaltose; D005964:Glucosyltransferases; C087517:cycloisomaltooligosaccharide glucanotransferase
keywords: carbohydrate binding modules; crystallography; cycloisomaltooligosaccharide; glycoside hydrolase