title: Characterization of a thermophilic 4-O-β-D-mannosyl-D-glucose phosphorylase from Rhodothermus marinus.
abstract: 4-O-β-D-Mannosyl-D-glucose phosphorylase (MGP), found in anaerobes, converts 4-O-β-D-mannosyl-D-glucose (Man-Glc) to α-D-mannosyl phosphate and D-glucose. It participates in mannan metabolism with cellobiose 2-epimerase (CE), which converts β-1,4-mannobiose to Man-Glc. A putative MGP gene is present in the genome of the thermophilic aerobe Rhodothermus marinus (Rm) upstream of the gene encoding CE. Konjac glucomannan enhanced production by R. marinus of MGP, CE, and extracellular mannan endo-1,4-β-mannosidase. Recombinant RmMGP catalyzed the phosphorolysis of Man-Glc through a sequential bi-bi mechanism involving ternary complex formation. Its molecular masses were 45 and 222 kDa under denaturing and nondenaturing conditions, respectively. Its pH and temperature optima were 6.5 and 75 °C, and it was stable between pH 5.5-8.3 and below 80 °C. In the reverse reaction, RmMGP had higher acceptor preferences for 6-deoxy-D-glucose and D-xylose than R. albus NE1 MGP. In contrast to R. albus NE1 MGP, RmMGP utilized methyl β-D-glucoside and 1,5-anhydro-D-glucitol as acceptor substrates.
authors.affiliation: a Research Faculty of Agriculture , Hokkaido University , Sapporo , Japan.
authors.forename: Nongluck
authors.initials: N
authors.lastname: Jaito
authors.forename: Wataru
authors.initials: W
authors.lastname: Saburi
authors.forename: Rei
authors.initials: R
authors.lastname: Odaka
authors.forename: Yusuke
authors.initials: Y
authors.lastname: Kido
authors.forename: Ken
authors.initials: K
authors.lastname: Hamura
authors.forename: Mamoru
authors.initials: M
authors.lastname: Nishimoto
authors.forename: Motomitsu
authors.initials: M
authors.lastname: Kitaoka
authors.forename: Hirokazu
authors.initials: H
authors.lastname: Matsui
authors.forename: Haruhide
authors.initials: H
authors.lastname: Mori
mesh_terms: D000595:Amino Acid Sequence; D004795:Enzyme Stability; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D008351:Mannans / Q000378:metabolism; D008364:Mannosyltransferases / Q000737:chemistry* / Q000378:metabolism*; D008969:Molecular Sequence Data; D010766:Phosphorylation; D045245:Rhodothermus / Q000201:enzymology*; D013379:Substrate Specificity; D013696:Temperature
chemical_list: D008351:Mannans; C022901:(1-6)-alpha-glucomannan; D008364:Mannosyltransferases
keywords: 4-O-β-d-mannosyl-d-glucose phosphorylase; Rhodothermus marinus; mannan; phosphorolysis; substrate specificity