Trehalose phosphorylase from Euglena gracilis

Belocopitow, Enrique; Maréchal, Luis R.

doi:10.1016/0005-2744(70)90045-8

Cited by 57 publications

(25 citation statements)

References 10 publications

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“…On the other hand, Belocopitow and Mar~chal (1970) reported phosphorolytic cleavage of trehalose into /9-GIc 1-P and glucose by a phosphorylase from a protist, Euglena gracilis Klebs. However, E-type trehalose phosphorylase was not detected in Homobasidiomycetes, Heterobasidiomycetes or Hemibasidiomycetes (Kitamoto et ah, 1998), a-Type trehalose phosphorylase is remarkably unstable, which makes it difficult to purify or estimate its weight described, as is the E-type trehalose phosphorylase by Mar6chal and Belocopitow (1972).…”

Section: Discussionmentioning

confidence: 94%

Purification and properties of α-glucose 1-phosphateforming trehalose phosphorylase from a basidiomycete, Pleurotus ostreatus

et al. 2000

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Pleurotus ostreatus produced a high activity of a-glucose 1-phosphate (a-GIc l-P) forming trehalose phosphorylase in vegetative mycelia and fruit-bodies, The enzyme was purified to homogeneity from the fruit-bodies by a procedure involving ammonium sulfate fractionation, DEAE-cellulose column chromatographies and cellulose phosphate column chromatographies. The enzyme catalyzes both the phosphorolysis of trehalose to produce a-GIc 1-P and glucose, and the synthesis of trehalose. It was not active toward other a-or ~-glucosyl disaccharides and polysaccharides. The optimum pH was 7.0 for phosphorolysis and 6.4 for synthesis of trehalose. The Km values for trehalose and Pi in phospholytic reaction were 76 mM and 4.2 raM, respectively. Those for glucose and a-GIc 1-P in synthetic reaction were 505 mM and 38 mM, respectively. The estimated molecular mass by the sedimentation equilibrium method using an ultracentrifuge was 120 kDa. The molecular mass of the subunit (61 kDa) by SDS-polyacrylamide gel electrophoresis suggested that the enzyme was a dimer of two identical subunitso The addition of glycerol higher than 25% into the enzyme solution stabilized its activity. The removal of phosphorus ions from the enzyme solution, by means of dialysis or electrophoresis, caused inactivation of the enzyme, probably by dissociation of the holoenzyme into the subunit proteins.

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Section: Discussionmentioning

confidence: 94%

Purification and properties of α-glucose 1-phosphateforming trehalose phosphorylase from a basidiomycete, Pleurotus ostreatus

et al. 2000

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“…The glucose formed enters the glycolysis by glucokinase while ␤-G1P is converted to glucose 6-phosphate by ␤-phosphoglucomutase (␤-PGM) before entering the glycolysis (29). ␤-PGM is repressed by glucose and lactose and induced by maltose and trehalose (28), suggesting that trehalose is also catabolized by a phosphorylase, analogous to observations in Euglena gracilis (2) and in Micrococcus varians (16). However, the initial catabolism of maltose and trehalose is still not well established in L. lactis, and the possibility of alternative pathways has not been investigated.…”

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confidence: 72%

Physiological Role of β-Phosphoglucomutase in Lactococcus lactis

Levander

Andersson

Rådström

2001

Appl Environ Microbiol

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A ␤-phosphoglucomutase (␤-PGM) mutant of Lactococcus lactis subsp. lactis ATCC 19435 was constructed using a minimal integration vector and double-crossover recombination. The mutant and the wild-type strain were grown under controlled conditions with different sugars to elucidate the role of ␤-PGM in carbohydrate catabolism and anabolism. The mutation did not significantly affect growth, product formation, or cell composition when glucose or lactose was used as the carbon source. With maltose or trehalose as the carbon source the wild-type strain had a maximum specific growth rate of 0.5 h ؊1 , while the deletion of ␤-PGM resulted in a maximum specific growth rate of 0.05 h ؊1 on maltose and no growth at all on trehalose. Growth of the mutant strain on maltose resulted in smaller amounts of lactate but more formate, acetate, and ethanol, and approximately 1/10 of the maltose was found as ␤-glucose 1-phosphate in the medium. Furthermore, the ␤-PGM mutant cells grown on maltose were considerably larger and accumulated polysaccharides which consisted of ␣-1,4-bound glucose units. When the cells were grown at a low dilution rate in a glucose and maltose mixture, the wild-type strain exhibited a higher carbohydrate content than when grown at higher growth rates, but still this content was lower than that in the ␤-PGM mutant. In addition, significant differences in the initial metabolism of maltose and trehalose were found, and cell extracts did not digest free trehalose but only trehalose 6-phosphate, which yielded ␤-glucose 1-phosphate and glucose 6-phosphate. This demonstrates the presence of a novel enzymatic pathway for trehalose different from that of maltose metabolism in L. lactis.In Lactococcus lactis maltose is split into glucose and ␤-glucose 1-phosphate (␤-G1P) by a P i -dependent reaction catalyzed by maltose phosphorylase (23). The glucose formed enters the glycolysis by glucokinase while ␤-G1P is converted to glucose 6-phosphate by ␤-phosphoglucomutase (␤-PGM) before entering the glycolysis (29). ␤-PGM is repressed by glucose and lactose and induced by maltose and trehalose (28), suggesting that trehalose is also catabolized by a phosphorylase, analogous to observations in Euglena gracilis (2) and in Micrococcus varians (16). However, the initial catabolism of maltose and trehalose is still not well established in L. lactis, and the possibility of alternative pathways has not been investigated.To our knowledge, maltose and trehalose phosphorylase, together with ␤-PGM, are the only known enzymes in any organism that catalyze reactions involving ␤-G1P. Although the presence of ␤-PGM and ␤-G1P in both bacteria and algae has been reported, information about the metabolic role of this enzyme and intermediate is scarce. There are, however, indications that ␤-G1P can be used as a precursor for cell wall material in L. lactis (31), and it has been found to be a component of glycan in Enterococcus faecalis (26).In this study, we constructed a ␤-PGM mutant of L. lactis which was used to further elucidate the rol...

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“…TPs have been found in various organisms such as Grifola frondosa (Saito et al, 1998), Schizophyllum commune (Eis & Nidetzky, 1999), Agaricus bisporus (Wannet et al, 1998), Catellatospora ferruginea (Aisaka & Masuda, 1995), Thermoanaerobacter brockii (Chaen et al, 1999) and Euglena gracilis (Belocopitow & Maré chal, 1970). The TPs above can be classified into two groups according to the anomeric specificity of d-glucose 1-phosphate.…”

Section: Introductionmentioning

confidence: 99%

Crystallization and X-ray diffraction studies of inverting trehalose phosphorylase fromThermoanaerobacter sp.

et al. 2010

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Disaccharide phosphorylases are attractive enzymatic platforms for tailor-made sugar synthesis owing to their ability to catalyze both the synthesis and the breakdown of disaccharides. Trehalose phosphorylase from Thermoanaerobacter sp. (TP) is a glycoside hydrolase family 65 enzyme which catalyzes the reversible breakdown of trehalose [d-glucopyranosyl-(1,1)-d-glucopyranose] to -d-glucose 1-phosphate and d-glucose. Recombinant purified protein was produced in Escherichia coli and crystallized in space group P2 1 2 1 2 1 . Crystals of recombinant TP were obtained in their native form and were soaked with glucose, with n-octyl--d-glucoside and with trehalose. The crystals presented a number of challenges including an unusually large unit cell, with a c axis measuring 420 Å , and variable diffraction quality. Crystal-dehydration protocols led to improvements in diffraction quality that were often dramatic, typically from 7-8 to 3-4 Å resolution. The structure of recombinant TP was determined by molecular replacement to 2.8 Å resolution, thus establishing a starting point for investigating the structural and mechanistic determinants of the disaccharide phosphorylase activity. To the best of our knowledge, this is the first crystal structure determination of an inverting trehalose phosphorylase.

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Trehalose phosphorylase from Euglena gracilis

Cited by 57 publications

References 10 publications

Purification and properties of α-glucose 1-phosphateforming trehalose phosphorylase from a basidiomycete, Pleurotus ostreatus

Purification and properties of α-glucose 1-phosphateforming trehalose phosphorylase from a basidiomycete, Pleurotus ostreatus

Physiological Role of β-Phosphoglucomutase in Lactococcus lactis

Crystallization and X-ray diffraction studies of inverting trehalose phosphorylase fromThermoanaerobacter sp.

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