1668-09-3Relevant articles and documents
Determination of kinetic parameters for maltotriose and higher malto-oligosaccharides in the reactions catalyzed by α-D-glucan phosphorylase from potato
Suganuma,Kitazono,Yoshinaga,Fujimoto,Nagahama
, p. 213 - 220 (1991)
For kinetic studies on its synthetic and phosphorolytic reactions, α-D-glucan phosphorylase from potatoes was purified chromatographically until free of D-enzyme. Purified maltotriose (G3) is a poor primer in the phosphorylase-catalyzed synthetic reaction, showing an anomalous time course and making previous attempts to determine its kinetic parameters unsuccessful. In the present work the true rate of the G3-primed reaction was obtained from linear plots obtained by incorporating a sufficient quantity of β-amylase in the digest to eliminate the more rapidly reacting G4 formed from the G3. A K(m) value of 9.4 ± 0.8 mM for G3 was calculated from the data by a nonlinear least-squares method. Kinetic parameters for a series of higher malto-oligosaccharides (G4-G8) were also determined in both the synthetic and the phosphorolytic directions. A large change in the values of K(m) and V/e was seen on going from G3 to G4 for the synthetic reaction, and from G4 to G5 for the phosphorolytic. For the higher saccharides the V/e values do not vary strongly with increasing d.p., while the K(m) values tend to decrease, as has seen in the reactions of other plant phosphorylases. For kinetic studies on its synthetic and phosphorolytic reactions α-D-glucan phosphorylase from potatoes was purified chromatographically until free of D-enzyme. Purified maltotriose (G3) is a poor primer in the phosphorylase-catalyzed synthetic reaction, showing an anomalous time course and making previous attempts to determine its kinetic parameters unsuccessful. In the present work the true rate of the G3-primed reaction was obtained from linear plots obtained by incorporating a sufficient quantity of β-amylase in the digest to eliminate the more rapidly reacting G4 formed from the G4 A Km value of 9.4 ± 0.8 mM for G3 was calculated from the data by a nonlinear least-squares method. Kinetic parameters for a series of higher malto-oligosaccharides (G4-G3) were also determined in both the synthetic and the phosphorolytic directions. A large change in the values of Km and V/e was seen on going from G3 to G4 for the synthetic reaction, and from G4 to G3 for the phosphorolytic. For the higher saccharides the V/e values do not vary strongly with increasing d.p.. while the Km values tend to decrease, as has seen in the reactions of other plant phosphorylases.
Automated Assembly of Starch and Glycogen Polysaccharides
Delbianco, Martina,Seeberger, Peter H.,Zhu, Yuntao
supporting information, p. 9758 - 9768 (2021/06/30)
Polysaccharides are Nature's most abundant biomaterials essential for plant cell wall construction and energy storage. Seemingly minor structural differences result in entirely different functions: cellulose, a β (1-4) linked glucose polymer, forms fibrils that can support large trees, while amylose, an α (1-4) linked glucose polymer forms soft hollow fibers used for energy storage. A detailed understanding of polysaccharide structures requires pure materials that cannot be isolated from natural sources. Automated Glycan Assembly provides quick access to trans-linked glycans analogues of cellulose, but the stereoselective installation of multiple cis-glycosidic linkages present in amylose has not been possible to date. Here, we identify thioglycoside building blocks with different protecting group patterns that, in concert with temperature and solvent control, achieve excellent stereoselectivity during the synthesis of linear and branched α-glucan polymers with up to 20 cis-glycosidic linkages. The molecules prepared with the new method will serve as probes to understand the biosynthesis and the structure of α-glucans.
Phosphorylase-catalyzed N-formyl-α-glucosaminylation of maltooligosaccharides
Kawazoe, Satoshi,Izawa, Hironori,Nawaji, Mutsuki,Kaneko, Yoshiro,Kadokawa, Jun-ichi
experimental part, p. 631 - 636 (2010/09/20)
This paper describes the phosphorylase-catalyzed enzymatic N-formyl-α-glucosaminylation of maltooligosaccharides for direct incorporation of 2-deoxy-2-formamido-α-d-glucopyranose units into maltooligosaccharides. When the reaction of 2-deoxy-2-formamido-α-d-glucopyranose-1-phosphate (GlcNF-1-P) as the glycosyl donor and maltotetraose as a glycosyl acceptor was performed in the presence of phosphorylase, the N-formyl-α-d-glucosaminylated pentasaccharide was produced, as confirmed by MALDI-TOF MS. Furthermore, the glucoamylase-catalyzed reaction of the crude products supported that the 2-deoxy-2-formamido-α-d-glucopyranoside unit was positioned at the non-reducing end of the pentasaccharide. The pentasaccharide was isolated from the crude products and its structure was further determined by the 1H NMR analysis. On the other hand, when the phosphorylase-catalyzed reactions of maltotriose and maltopentaose using GlcNF-1-P were conducted, no N-formyl-α-glucosaminylation took place in the former system, whereas the latter system gave N-formyl-α-d-glucosaminylated oligosaccharides with various degrees of polymerization. These results could be explained by the recognition behavior of phosphorylase toward maltooligosaccharides.
Expression, purification, and characterization of the maltooligosyltrehalose trehalohydrolase from the thermophilic archaeon Sulfolobus solfataricus ATCC 35092
Fang, Tsuei-Yun,Tseng, Wen-Chi,Guo, Meng-Shin,Shih, Tong-Yuan,Hung, Xing-Guang
, p. 7105 - 7112 (2008/02/03)
The maltooligosyltrehalose trehalohydrolase (MTHase) mainly cleaves the α-1,4-glucosidic linkage next to the α-1,1-linked terminal disaccharide of maltooligosyltrehalose to produce trehalose and the maltooligosaccharide with lower molecular mass. In this study, the treZ gene encoding MTHase was PCR-cloned from Sulfolobus solfataricus ATCC 35092 and then expressed in Escherichia coli. A high yield of the active wild-type MTHase, 13300 units/g of wet cells, was obtained in the absence of IPTG induction. Wild-type MTHase was purified sequentially using heat treatment, nucleic acid precipitation, and ion-exchange chromatography. The purified wild-type MTHase showed an apparent optimal pH of 5 and an optimal temperature at 85°C. The enzyme was stable at pH values ranging from 3.5 to 11, and the activity was fully retained after a 2-h incubation at 45-85°C. The kcat values of the enzyme for hydrolysis of maltooligosyltrehaloses with degree of polymerization (DP) 4-7 were 193, 1030, 1190, and 1230 s-1, respectively, whereas the kcat values for glucose formation during hydrolysis of DP 4-7 maltooligosaccharides were 5.49, 17.7, 18.2, and 6.01 s-1, respectively. The KM values of the enzyme for hydrolysis of DP 4-7 maltooligosyltrehaloses and those for maltooligosaccharides are similar at the same corresponding DPs. These results suggest that this MTHase could be used to produce trehalose at high temperatures.
