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Literature summary extracted from

  • Guerin, F.; Barbe, S.; Pizzut-Serin, S.; Potocki-Veronese, G.; Guieysse, D.; Guillet, V.; Monsan, P.; Mourey, L.; Remaud-Simeon, M.; Andre, I.; Tranier, S.
    Structural investigation of the thermostability and product specificity of amylosucrase from the bacterium Deinococcus geothermalis (2012), J. Biol. Chem., 287, 6642-6654.
    View publication on PubMedView publication on EuropePMC

Cloned(Commentary)

EC Number Cloned (Comment) Organism
2.4.1.4 expression of GST-tagged DgAS Deinococcus geothermalis
2.4.1.4 expression of GST-tagged DgAS Neisseria polysaccharea

Crystallization (Commentary)

EC Number Crystallization (Comment) Organism
2.4.1.4 purified recombinant detagged DgAS, free and in complex with turanose, hanging drop vapor diffusion method, X-ray diffraction structure determination and analysis at 1.97-2.10 A resolution Deinococcus geothermalis
2.4.1.4 purified recombinant detagged NpAS in complex with turanose, hanging drop vapor diffusion method, 1:1 v/v ratio of protein, containing 6 mg/ml in 20 mM Tris, pH 8.0, to precipitant solution containing 1.5 M sodium acetate, 0.1 M sodium cacodylate, pH 7.0, 2 weeks, X-ray diffraction structure determination and analysis at 1.85 A resolution Neisseria polysaccharea

Protein Variants

EC Number Protein Variants Comment Organism
2.4.1.4 E328Q inactive mutant Neisseria polysaccharea

Natural Substrates/ Products (Substrates)

EC Number Natural Substrates Organism Comment (Nat. Sub.) Natural Products Comment (Nat. Pro.) Rev. Reac.
2.4.1.4 sucrose + [(1->4)-alpha-D-glucosyl]n Deinococcus geothermalis
-
D-fructose + [(1->4)-alpha-D-glucosyl]n+1
-
?
2.4.1.4 sucrose + [(1->4)-alpha-D-glucosyl]n Neisseria polysaccharea
-
D-fructose + [(1->4)-alpha-D-glucosyl]n+1
-
?

Organism

EC Number Organism UniProt Comment Textmining
2.4.1.4 Deinococcus geothermalis
-
-
-
2.4.1.4 Neisseria polysaccharea Q9ZEU2
-
-

Purification (Commentary)

EC Number Purification (Comment) Organism
2.4.1.4 recombinant GST-tagged DgAS by glutathione affinity chromatography, followed by proteolytic removal of the GST-tag Deinococcus geothermalis
2.4.1.4 recombinant GST-tagged NpAS by glutathione affinity chromatography, followed by proteolytic removal of the GST-tag Neisseria polysaccharea

Reaction

EC Number Reaction Comment Organism Reaction ID
2.4.1.4 sucrose + [(1->4)-alpha-D-glucosyl]n = D-fructose + [(1->4)-alpha-D-glucosyl]n+1 DgAS binds the furanoid tautomers of fructose through a weak network of interactions to enable turanose formation. Such topology at subsite +1 is likely favoring other possible fructose binding modes as reported for Neisseria polysaccharea NpAS in agreement with the higher amount of trehalulose formed by DgAS. Residues Glu326 and Asp284 of DgAS are the general acid/base and the nucleophile, respectively, involved in the formation of the beta-glucosyl intermediate occurring in the alpha-retaining mechanism Deinococcus geothermalis
2.4.1.4 sucrose + [(1->4)-alpha-D-glucosyl]n = D-fructose + [(1->4)-alpha-D-glucosyl]n+1 in Neisseria polysaccharea NpAS key residues force the fructosyl moiety to bind in an open state with the O3' ideally positioned to explain the preferential formation of turanose by NpAS. Residues Glu328 and Asp286 of NpAS are the general acid/base and the nucleophile, respectively, involved in the formation of the beta-glucosyl intermediate occurring in the alpha-retaining mechanism Neisseria polysaccharea

Substrates and Products (Substrate)

EC Number Substrates Comment Substrates Organism Products Comment (Products) Rev. Reac.
2.4.1.4 additional information synthesis of sucrose isomers turanose and trehalulose from sucrose in the presence of fructose by DgAS, turanose binding site structure, overview Deinococcus geothermalis ?
-
?
2.4.1.4 additional information synthesis of sucrose isomers turanose and trehalulose from sucrose in the presence of fructose by NpAS, turanose binding site structure, overview Neisseria polysaccharea ?
-
?
2.4.1.4 sucrose + [(1->4)-alpha-D-glucosyl]n
-
Deinococcus geothermalis D-fructose + [(1->4)-alpha-D-glucosyl]n+1
-
?
2.4.1.4 sucrose + [(1->4)-alpha-D-glucosyl]n
-
Neisseria polysaccharea D-fructose + [(1->4)-alpha-D-glucosyl]n+1
-
?

Subunits

EC Number Subunits Comment Organism
2.4.1.4 homodimer three-dimensional structure and dimer interface analysis. The quaternary organization is likely to participate in the enhanced thermal stability of the protein. Structure comparison with the amylosucrase from Neisseria polysaccharea, overview Deinococcus geothermalis
2.4.1.4 More three-dimensional structure analysis. Structure comparison with the amylosucrase from Deinococcus geothermalis, overview Neisseria polysaccharea

Synonyms

EC Number Synonyms Comment Organism
2.4.1.4 DGAS
-
Deinococcus geothermalis
2.4.1.4 NPAS
-
Neisseria polysaccharea

Temperature Optimum [°C]

EC Number Temperature Optimum [°C] Temperature Optimum Maximum [°C] Comment Organism
2.4.1.4 50
-
sucrose isomer synthesis Deinococcus geothermalis
2.4.1.4 50
-
sucrose isomer synthesis Neisseria polysaccharea

Temperature Stability [°C]

EC Number Temperature Stability Minimum [°C] Temperature Stability Maximum [°C] Comment Organism
2.4.1.4 additional information
-
the quaternary organization is likely to participate in the enhanced thermal stability of the protein Deinococcus geothermalis
2.4.1.4 additional information
-
the quaternary organization is likely to participate in the enhanced thermal stability of the protein Neisseria polysaccharea

pH Optimum

EC Number pH Optimum Minimum pH Optimum Maximum Comment Organism
2.4.1.4 7
-
assay at Deinococcus geothermalis
2.4.1.4 7
-
assay at Neisseria polysaccharea

General Information

EC Number General Information Comment Organism
2.4.1.4 evolution the enzyme belongs to the glycohydrolase family 13, GH13 Deinococcus geothermalis
2.4.1.4 evolution the enzyme belongs to the glycohydrolase family 13, GH13 Neisseria polysaccharea
2.4.1.4 additional information the catalytic domain A of DgAS adopts the typical (alpha/alpha)8-barrel of the GH family 13 Deinococcus geothermalis
2.4.1.4 additional information the catalytic domain A of NpAS adopts the typical (alpha/alpha)8-barrel of the GH family 13 Neisseria polysaccharea
2.4.1.4 physiological function amylosucrases are sucrose-utilizing alpha-transglucosidases that naturally catalyze the synthesis of alpha-glucans, linked exclusively through alpha-1,4-linkages. Side products and in particular sucrose isomers such as turanose and trehalulose are also produced by these enzymes Deinococcus geothermalis
2.4.1.4 physiological function amylosucrases are sucrose-utilizing alpha-transglucosidases that naturally catalyze the synthesis of alpha-glucans, linked exclusively through alpha-1,4-linkages. Side products and in particular sucrose isomers such as turanose and trehalulose are also produced by these enzymes Neisseria polysaccharea