Subsite structure and catalytic mechanism of a new glycosyltrehalose-producing enzyme isolated from the hyperthermophilic archaeum, Sulfolobus solfataricus KM1.
A glycosyltrehalose-producing enzyme from Sulfolobus solfataricus KM1 catalyzes a conversion of maltooligosaccharides to glycosyltrehaloses and also hydrolyzes maltooligosaccharides to liberate glucose, as a side reaction. From the sum of the conversion and hydrolysis reaction rates, the rate parameters involved in the "splitting" of the alpha-1,4 glucosidic linkage were calculated. From the data obtained, the subsite structure for maltooligosaccharides was identified. From the analysis of the hydrolysate of maltotriose in [18O labeled H2O, the hypothesis of the C1-O bond splitting and the formation of a glycosyl (maltosyl)-enzyme intermediate was strongly supported. From the analysis of the reaction product in the presence of [3H] labeled glucose, the occurrence of intermolecular transglycosylation was confirmed. These data strongly support the suggestion that the catalytic mechanism of this enzyme is a transglycosylation.
Kato
M
M
Applied Bioresearch Center, Kirin Brewery Co., Ltd. Gunma, Japan. kmasaru@kirin.co.jp
Takehara
K
K
Kettoku
M
M
Kobayashi
K
K
Shimizu
T
T
D002384:Catalysis; D005964:Glucosyltransferases / Q000737:chemistry* / Q000378:metabolism*; D007700:Kinetics; D011487:Protein Conformation; D016339:Spectrometry, Mass, Fast Atom Bombardment; D013379:Substrate Specificity; D016944:Sulfolobus / Q000201:enzymology*
D005964:Glucosyltransferases; C408377:glycosyltransferase, Sulfolobus