Enzymatic characterization and substrate specificity of thermostable beta-glycosidase from hyperthermophilic archaea, Sulfolobus shibatae, expressed in E. coli.
Enzymatic properties and substrate specificity of recombinant beta-glycosidases from a hyperthermophilic archaeon, Sulfolobus shibatae (rSSG), were analyzed. rSSG showed its optimum temperature and pH at 95 degrees C and pH 5.0, respectively. Thermal inactivation of rSSG showed that its half-life of enzymatic activity at 75 degrees C was 15 h whereas it drastically decreased to 3.9 min at 95 degrees C. The addition of 10 mM of MnCl2 enhanced the hydrolysis activity of rSSG up to 23% whereas most metal ions did not show any considerable effect. Dithiothreitol (DTT) and 2-mercaptoethanol exhibited significant influence on the increase of the hydrolysis activity of rSSG. rSSG apparently preferred laminaribiose (beta1-->3Glc), followed by sophorose (beta1-->2Glc), gentiobiose (beta1-->6Glc), and cellobiose (beta1--4Glc). Various intermolecular transfer products were formed by rSSG in the lactose reaction, indicating that rSSG prefers lactose as a good acceptor as well as a donor. The strong intermolecular transglycosylation activity of rSSG can be applied in making functional oligosaccharides.
Park
Na-Young
NY
Department of Food Science and Biotechnology, Graduate School of Biotechnology & Institute of Life Sciences and Resources, KyungHee University, Yongin 449-701, Korea.
Cha
Jaeho
J
Kim
Dae-Ok
DO
Park
Cheon-Seok
CS
D001709:Biotechnology; D002240:Carbohydrate Sequence; D019281:Dimerization; D004795:Enzyme Stability; D004926:Escherichia coli / Q000201:enzymology / Q000235:genetics; D005959:Glucosidases / Q000235:genetics / Q000378:metabolism*; D006031:Glycosylation; D006358:Hot Temperature; D006863:Hydrogen-Ion Concentration; D008969:Molecular Sequence Data; D009844:Oligosaccharides / Q000737:chemistry / Q000378:metabolism; D013379:Substrate Specificity; D016944:Sulfolobus / Q000201:enzymology* / Q000235:genetics; D013696:Temperature
D009844:Oligosaccharides; D005959:Glucosidases; C104851:Sulfolobus solfataricus beta-glycosidase