Enzymatic synthesis of a 2-O-α-d-glucopyranosyl cyclic tetrasaccharide by kojibiose phosphorylase

Watanabe, Hikaru; Higashiyama, Takanobu; Aga, Hajime; Nishimoto, Tetsuya; Kubota, Michio; Fukuda, Shigeharu; Kurimoto, Masashi; Tsujisaka, Yoshio

doi:10.1016/j.carres.2004.12.012

Cited by 16 publications

(3 citation statements)

References 23 publications

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“…In literature, the synthesis of α-glucose-1-phosphate (αGlc1P) from starch [10,11] or sucrose [12] and that of α-galactose-1-phosphate (αGal1P) from lactose [13,14] have previously been reported. A production process for βGlc1P has not yet been described in detail, but is similarly feasible using GH65 disaccharide phosphorylases from cheap substrates [15] briefly mention the production of βGlc1P from trehalose, using typical reaction conditions employed for αGlc1P [16]. In our hands, however, a crystalline product could not be obtained using these procedures.…”

Section: Introductionmentioning

confidence: 95%

Enzymatic production of β‐D‐glucose‐1‐phosphate from trehalose

Borght¹,

Desmet²,

Soetaert³

2010

Biotechnology Journal

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beta-D-Glucose-1-phosphate (beta Glc1P) is an efficient glucosyl donor for both enzymatic and chemical glycosylation reactions but is currently very costly and not available in large amounts. This article provides an efficient production method of beta Glc1P from trehalose and phosphate using the thermostable trehalose phosphorylase from Thermoanaerobacter brockii. At the process temperature of 60 C, Escherichia coli expression host cells are lysed and cell treatment prior to the reaction is, therefore, not required. In this way, the theoretical maximum yield of 26% could be easily achieved. Two different purification strategies have been compared, anion exchange chromatography or carbohydrate removal by treatment with trehalase and yeast, followed by chemical phosphate precipitation. In a next step, beta Glc1P was precipitated with ethanol but this did not induce crystallization, in contrast to what ist observed with other glycosylphosphates. After conversion of the product to its cyclohexylammonium salt, however, crystals could be readily obtained. Although both purification methods were quantitative (>99% recovery), a large amount of product (50%) was lost during crystallization. Nevertheless, a production process for crystalline beta Glc1P is now available from the cheap substrates trehalose and inorganic phosphate

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Section: Introductionmentioning

confidence: 95%

Enzymatic production of β‐D‐glucose‐1‐phosphate from trehalose

Borght¹,

Desmet²,

Soetaert³

2010

Biotechnology Journal

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“…Beyond simpler sugar acceptors, this enzyme could be used for the addition of d -glucose to a cyclic α- d -glucan, forming cyclo -{ d -Glc-α-1,3- d -Glc-[α-1,2- d -Glc-]-α-1,6- d -Glc-α-1,3- d -Glc p -α-1,6-} ( 26 ). 29 This enzyme was also noted as transferring d -glucose onto l -sorbose and l -xylose, though it is unclear to which hydroxyl group the glycosidic bond is formed. 25 …”

Section: α-12- D -Glucan Phosphorylasesmentioning

confidence: 99%

Enzymatic synthesis using glycoside phosphorylases

O’Neill

Field

2015

Carbohydrate Research

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“…KP activity was first identified from Thermoanaerobacter brockii , ATCC 35047 . The properties and synthetic abilities of T. brockii KP (TbKP) have been studied in detail , and it has been shown that TbKP can synthesize kojibiose and other oligosaccharides containing α1,2‐linked glucose residues. Recently, KP from Caldicellulosiruptor saccharolyticus , ATCC 43494/DSM 8903 (CsKP; Csac_0439, 55% sequence identity with TbKP), which has higher thermostability and wider substrate specificity than TbKP, has been cloned and characterized .…”

Section: Introductionmentioning

confidence: 99%

Structural and mutational analysis of substrate recognition in kojibiose phosphorylase

et al. 2013

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Glycoside hydrolase (GH) family 65 contains phosphorylases acting on maltose (Glc-a1,4-Glc), kojibiose (Glc-a1,2-Glc), trehalose (Glc-a1,a1,-Glc), and nigerose (Glc-a1,3-Glc). These phosphorylases can efficiently catalyze the reverse reactions with high specificities, and thus can be applied to the practical synthesis of a-glucosyl oligosaccharides. Here, we determined the crystal structures of kojibiose phosphorylase from Caldicellulosiruptor saccharolyticus in complex with glucose and phosphate and in complex with kojibiose and sulfate, providing the first structural insights into the substrate recognition of a glycoside hydrolase family 65 enzyme. The loop 3 region comprising the active site of kojibiose phosphorylase is significantly longer than the active sites of other enzymes, and three residues around this loop, Trp391, Glu392, and Thr417, recognize kojibiose. Various mutants mimicking the residue conservation patterns of other phosphorylases were constructed by mutation at these three residues. Activity measurements of the mutants against four substrates indicated that Trp391 and Glu392, especially the latter, are required for the kojibiose activity.

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Enzymatic synthesis of a 2-O-α-d-glucopyranosyl cyclic tetrasaccharide by kojibiose phosphorylase

Cited by 16 publications

References 23 publications

Enzymatic production of β‐D‐glucose‐1‐phosphate from trehalose

Enzymatic production of β‐D‐glucose‐1‐phosphate from trehalose

Enzymatic synthesis using glycoside phosphorylases

Structural and mutational analysis of substrate recognition in kojibiose phosphorylase

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