title: Cloning, expression and characterization of a thermostable exo-beta-D-glucosaminidase from the hyperthermophilic archaeon Pyrococcus horikoshii.
abstract: An exo-beta-D-glucosaminidase gene (PH0511) was cloned from the hyperthermophilic archaeon, Pyrococcus horikoshii, and expressed in Escherichia coli. The purified protein showed a strong exo-beta-D: -glucosaminidase activity by TLC analysis. DTT (50 mM) had little effect on its homodimeric structure during SDS-PAGE. The enzyme was optimally active at 90 degrees C (over 20 min) and pH 6. It had a half-life of 9 h at 90 degrees C and is the most thermostable glucosaminidase described up to now. The activity was not inhibited by ethanol, 2-propanol, DMSO, PEG-400, denaturing agents SDS (5%, w/v), urea, guanidine hydrochloride (5 M) and Mg(2+), Mn(2+), Co(2+), Ca(2+), Sr(2+), Ni(2+) (at up to 10 mM).
authors.affiliation: State Key Laboratory of Microbial Technology, University of Shandong, 250100, Shandong, Jinan, P.R. China.
authors.forename: Bo
authors.initials: B
authors.lastname: Liu
authors.forename: Zhuo
authors.initials: Z
authors.lastname: Li
authors.forename: Ye
authors.initials: Y
authors.lastname: Hong
authors.forename: Jinfeng
authors.initials: J
authors.lastname: Ni
authors.forename: Duohong
authors.initials: D
authors.lastname: Sheng
authors.forename: Yulong
authors.initials: Y
authors.lastname: Shen
mesh_terms: D002413:Cations, Divalent; D003001:Cloning, Molecular; D019281:Dimerization; D004121:Dimethyl Sulfoxide / Q000737:chemistry; D004229:Dithiothreitol / Q000737:chemistry; D004795:Enzyme Stability; D004926:Escherichia coli / Q000378:metabolism; D019791:Guanidine / Q000737:chemistry; D006596:Hexosaminidases / Q000096:biosynthesis* / Q000737:chemistry / Q000302:isolation & purification; D006863:Hydrogen-Ion Concentration; D043902:Pyrococcus horikoshii / Q000201:enzymology*; D011994:Recombinant Proteins / Q000037:antagonists & inhibitors / Q000096:biosynthesis / Q000302:isolation & purification; D012967:Sodium Dodecyl Sulfate / Q000737:chemistry; D013696:Temperature; D014508:Urea / Q000737:chemistry
chemical_list: D002413:Cations, Divalent; D011994:Recombinant Proteins; D012967:Sodium Dodecyl Sulfate; D014508:Urea; D006596:Hexosaminidases; C064499:exo-beta-D-glucosaminidase; D019791:Guanidine; D004229:Dithiothreitol; D004121:Dimethyl Sulfoxide