Facile synthesis of glucose-1-phosphate from starch by Thermus caldophilus GK24 α-glucan phosphorylase

Bae, Jungdon; Lee, DuckHee; Kim, Doo-Il; Cho, Soo-Jin; Park, Jung Eun; Koh, Sukhoon; Kim, Joong-Su; Park, Bo-Hyun; Choi, Yongseok; Shin, Hyun‐Jae; Hong, Seok-In; Lee, Dae-Sil

doi:10.1016/j.procbio.2005.05.007

Cited by 14 publications

(14 citation statements)

References 21 publications

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“…Glycoside phosphorylases, in contrast, only need inorganic phosphate to produce a glycosylphosphate. 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].…”

Section: Introductionmentioning

confidence: 99%

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: 99%

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

Borght¹,

Desmet²,

Soetaert³

2010

Biotechnology Journal

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“…It has been observed that most known phosphorylases have a subunit molecular mass of around 90 kDa and usually exist as homodimers or tetramers (Palm et al 1985). At 25°C the thermostable α-glucan phosphorylases from Thermus thermophilus, Thermus caldophilus and Thermococcus litoralis form octamers, hexamers and trimers, respectively (Boeck and Schinzel 1996, Xavier et al 1999, Bae et al 2005.…”

Section: Cloning and Purification Of The Pf1535 Proteinmentioning

confidence: 99%

“…Significant progress has been made with the detailed structural and functional characterization of glycogen phosphorylases of mesophilic origins (Chen and Segel 1968, Hwang and Fletterick 1986, Browner et al 1991, Rogers et al 1992, Srivastava et al 1996; however, little is known about those from hyperthermophilic organisms. Thermostable α-glucan phosphorylases reported to date have been isolated from the thermophilic bacteria Bacillus stearothermophilus, Thermus caldophilus, Thermus aquaticus, Thermus thermophilus, Thermotoga maritima and Aquifex aeolicus (Konig et al 1982, Boeck and Schinzel 1996, Bibel et al 1998, Takata et al 1998, Takaha et al 2001, Bhuiyan et al 2003, Bae et al 2005. Among archaea, the enzyme activity has been detected in Methanococcus maripaludis and Thermococcus litoralis (Yu et al 1994, Xavier et al 1999.…”

Section: Introductionmentioning

confidence: 99%

“…Glucan phosphorylases are valuable for the synthesis of G-1-P (Shin et al 2000, Bae et al 2005), a potential antibiotic or immunosuppressive drug (Takata et al 1998), for the production of linear amylose and other related glucose polymers (Yanase et al 2002, Fujii et al 2003, Ohdan et al 2007, for the synthesis of amylose-lipid complexes (Gelders et al 2005) and for carbohydrate engineering (Yanase et al 2006). The hyperthermostable glucan phosphorylase in combination with other thermostable sugar nucleotidyltransferases (Mizanur et al 2004(Mizanur et al , 2005 in a reaction using starch as the starting polysaccharide for the subsequent synthesis of nucleotide sugars could be advantageous over the mesophilic counterpart, because the solubility of starch is increased at higher temperature.…”

Section: Introductionmentioning

confidence: 99%

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Recombinant production and biochemical characterization of a hyperthermostable α‐glucan/maltodextrin phosphorylase from Pyrococcus furiosus

Rahman

Griffin

Pohl

2007

Archaea

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Alpha-glucan phosphorylase catalyzes the reversible cleavage of alpha-1-4-linked glucose polymers into alpha-D-glucose-1-phosphate. We report the recombinant production of an alpha-glucan/maltodextrin phosphorylase (PF1535) from a hyperthermophilic archaeon, Pyrococcus furiosus, and the first detailed biochemical characterization of this enzyme from any archaeal source using a mass-spectrometry-based assay. The apparent 98 kDa recombinant enzyme was active over a broad range of temperatures and pH, with optimal activity at 80 degrees C and pH 6.5-7. This archaeal protein retained its complete activity after 24 h at 80 degrees C in Tris-HCl buffer. Unlike other previously reported phosphorylases, the Ni-affinity column purified enzyme showed broad substrate specificity in both the synthesis and degradation of maltooligosaccharides. In the synthetic direction of the enzymatic reaction, the lowest oligosaccharide required for the chain elongation was maltose. In the degradative direction, the archaeal enzyme can produce glucose-1-phosphate from maltotriose or longer maltooligosaccharides including both glycogen and starch. The specific activity of the enzyme at 80 degrees C in the presence of 10 mM maltoheptaose and at 10 mg ml(-1) glycogen concentration was 52 U mg(-1) and 31 U mg(-1), respectively. The apparent Michaelis constant and maximum velocity for inorganic phosphate were 31 +/- 2 mM and 0.60 +/- 0.02 mM min(-1) microg(-1), respectively. An initial velocity study of the enzymatic reaction indicated a sequential bi-bi catalytic mechanism. Unlike the more widely studied mammalian glycogen phosphorylase, the Pyrococcus enzyme is active in the absence of added AMP.

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“…In the case of recombinant E. coli MP, a removal of trace phosphatase amounts cleaving Glc1P was essential and this could be achieved neither by aYnity precipitation with starch, or precipitation with Bioprocessing Agent BAP 1050 (Toso Haas, Stuttgart, Germany), nor by ion-exchange chromatography (Q-Sepharose, Pharmacia) or anion-exchange membrane Wltration (CIDE 1000, CIQM 1000, Millipore) [3]. Later, thermostable MPs from Thermus caldophilus [4,5] and Thermoanaerobacter tengcongensis [6] were explored for Glc1P production. In addition to the obvious potential advantage of higher process temperature to promote better solubilization and disorganization of the raw materials [7], thermostable enzymes can be overexpressed in a mesophilic host whose enzymes are completely inactivated during the thermal reaction.…”

Section: Introductionmentioning

confidence: 99%

Physiological aggregation of maltodextrin phosphorylase from Pyrococcus furiosus and its application in a process of batch starch degradation to α-d-glucose-1-phosphate

Nahálka

2007

J Ind Microbiol Biotechnol

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Maltodextrin phosphorylase from Pyrococcus furiosus (PF1535) was fused with the cellulose-binding domain of Clostridium cellulovorans serving as an aggregation module. After molecular cloning of the corresponding gene fusion construct and controlled expression in Escherichia coli BL21, 83% of total maltodextrin phosphorylase activity (0.24 U/mg of dry cell weight) was displayed in active inclusion bodies. These active inclusion bodies were easily isolated by nonionic detergent treatment and directly used for maltodextrin conversion to alpha-D-glucose-1-phosphate in a repetitive batch mode. Only 10% of enzyme activity was lost after ten conversion cycles at optimum conditions.

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Facile synthesis of glucose-1-phosphate from starch by Thermus caldophilus GK24 α-glucan phosphorylase

Cited by 14 publications

References 21 publications

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

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

Recombinant production and biochemical characterization of a hyperthermostable α‐glucan/maltodextrin phosphorylase from Pyrococcus furiosus

Physiological aggregation of maltodextrin phosphorylase from Pyrococcus furiosus and its application in a process of batch starch degradation to α-d-glucose-1-phosphate

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