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Literature summary extracted from

  • Mardo, K.; Visnapuu, T.; Gromkova, M.; Aasamets, A.; Viigand, K.; Vija, H.; Alamaee, T.
    High-throughput assay of levansucrase variants in search of feasible catalysts for the synthesis of fructooligosaccharides and levan (2014), Molecules, 19, 8434-8455.
    View publication on PubMedView publication on EuropePMC

Application

EC Number Application Comment Organism
2.4.1.10 synthesis levansucrase Lsc3 of Pseudomonas syringae pv. tomato has very high catalytic activity and stability making it a promising biotechnological catalyst for FOS and levan synthesis Pseudomonas syringae pv. tomato

Cloned(Commentary)

EC Number Cloned (Comment) Organism
2.4.1.10 gene lsc3, sequence comparison, recombinant expression of N-terminally His6-tagged wild-type and mutant enzymes in Escherichia coli strain BL21(DE3) Pseudomonas syringae pv. tomato

Protein Variants

EC Number Protein Variants Comment Organism
2.4.1.10 D219A site-directed mutagenesis, inactive mutant Pseudomonas syringae pv. tomato
2.4.1.10 D225A site-directed mutagenesis, the mutant shows reduced activity compared to the wild-type enzyme Pseudomonas syringae pv. tomato
2.4.1.10 D225N site-directed mutagenesis, the mutant shows enhanced activity compared to the wild-type enzyme Pseudomonas syringae pv. tomato
2.4.1.10 D300A site-directed mutagenesis, the mutant shows slightly enhanced activity compared to the wild-type enzyme Pseudomonas syringae pv. tomato
2.4.1.10 D333A site-directed mutagenesis, the mutant shows reduced activity compared to the wild-type enzyme Pseudomonas syringae pv. tomato
2.4.1.10 D333N site-directed mutagenesis, the mutant shows reduced activity compared to the wild-type enzyme Pseudomonas syringae pv. tomato
2.4.1.10 D62A site-directed mutagenesis, inactive mutant Pseudomonas syringae pv. tomato
2.4.1.10 E211Q site-directed mutagenesis, the mutant shows highly reduced activity compared to the wild-type enzyme Pseudomonas syringae pv. tomato
2.4.1.10 E236Q site-directed mutagenesis, the mutant shows highly reduced activity compared to the wild-type enzyme Pseudomonas syringae pv. tomato
2.4.1.10 E303A site-directed mutagenesis, inactive mutant Pseudomonas syringae pv. tomato
2.4.1.10 E303Q site-directed mutagenesis, inactive mutant Pseudomonas syringae pv. tomato
2.4.1.10 H113A site-directed mutagenesis, the mutant shows highly reduced activity compared to the wild-type enzyme Pseudomonas syringae pv. tomato
2.4.1.10 H113Q site-directed mutagenesis, inactive mutant Pseudomonas syringae pv. tomato
2.4.1.10 H306A site-directed mutagenesis, the mutant shows reduced activity compared to the wild-type enzyme Pseudomonas syringae pv. tomato
2.4.1.10 H321K site-directed mutagenesis, almost inactive mutant Pseudomonas syringae pv. tomato
2.4.1.10 H321L site-directed mutagenesis, almost inactive mutant Pseudomonas syringae pv. tomato
2.4.1.10 H321R site-directed mutagenesis, almost inactive mutant Pseudomonas syringae pv. tomato
2.4.1.10 H321R site-directed mutagenesis, significantly decreased polymerizing ability compared to the wild-type Pseudomonas syringae pv. tomato
2.4.1.10 H321S site-directed mutagenesis, almost inactive mutant Pseudomonas syringae pv. tomato
2.4.1.10 L66A site-directed mutagenesis, the mutant shows highly reduced activity compared to the wild-type enzyme Pseudomonas syringae pv. tomato
2.4.1.10 additional information Vmax values for sucrose splitting of mutants, structure-function analysis of enzyme mutants, overview Pseudomonas syringae pv. tomato
2.4.1.10 P220A site-directed mutagenesis, the mutant shows reduced activity compared to the wild-type enzyme Pseudomonas syringae pv. tomato
2.4.1.10 Q301A site-directed mutagenesis, significantly decreased polymerizing ability compared to the wild-type Pseudomonas syringae pv. tomato
2.4.1.10 Q301E site-directed mutagenesis, the mutant shows slightly enhanced activity compared to the wild-type enzyme Pseudomonas syringae pv. tomato
2.4.1.10 R304A site-directed mutagenesis, inactive mutant Pseudomonas syringae pv. tomato
2.4.1.10 R304C site-directed mutagenesis, inactive mutant Pseudomonas syringae pv. tomato
2.4.1.10 T302M site-directed mutagenesis, the mutant shows slightly reduced activity compared to the wild-type enzyme Pseudomonas syringae pv. tomato
2.4.1.10 T302P site-directed mutagenesis, the mutant shows reduced activity compared to the wild-type enzyme Pseudomonas syringae pv. tomato
2.4.1.10 T302P site-directed mutagenesis, significantly decreased polymerizing ability compared to the wild-type Pseudomonas syringae pv. tomato
2.4.1.10 W109A site-directed mutagenesis, almost inactive mutant Pseudomonas syringae pv. tomato
2.4.1.10 W61A site-directed mutagenesis, inactive mutant Pseudomonas syringae pv. tomato
2.4.1.10 W61N site-directed mutagenesis, inactive mutant Pseudomonas syringae pv. tomato
2.4.1.10 W80R site-directed mutagenesis, the mutant shows highly reduced activity compared to the wild-type enzyme Pseudomonas syringae pv. tomato

Localization

EC Number Localization Comment Organism GeneOntology No. Textmining
2.4.1.10 extracellular
-
 Pseudomonas syringae pv. tomato
-
-

Natural Substrates/ Products (Substrates)

EC Number Natural Substrates Organism Comment (Nat. Sub.) Natural Products Comment (Nat. Pro.) Rev. Reac.
2.4.1.10 sucrose + alpha-D-glucosyl-(1->2)-[(2->6)-alpha-D-fructosyl]n Pseudomonas syringae pv. tomato
-
 D-glucose + alpha-D-glucosyl-(1->2)-[(2->6)-alpha-D-fructosyl]n+1
-
 ?
2.4.1.10 sucrose + alpha-D-glucosyl-(1->2)-[(2->6)-alpha-D-fructosyl]n Pseudomonas syringae pv. tomato DC3000
-
 D-glucose + alpha-D-glucosyl-(1->2)-[(2->6)-alpha-D-fructosyl]n+1
-
 ?

Organism

EC Number Organism UniProt Comment Textmining
2.4.1.10 Pseudomonas syringae pv. tomato Q88BN6 pv. tomato, gene lsc3
-
2.4.1.10 Pseudomonas syringae pv. tomato DC3000 Q88BN6 pv. tomato, gene lsc3
-

Specific Activity [micromol/min/mg]

EC Number Specific Activity Minimum [µmol/min/mg] Specific Activity Maximum [µmol/min/mg] Comment Organism
2.4.1.10 additional information
-
 transfructosylation activity of wild-type and mutant enzymes (permeabilized cells/purified protein), overview Pseudomonas syringae pv. tomato
2.4.1.10 461.6
-
 purified recombinant wild-type enzyme, pH 6.0, 37°C Pseudomonas syringae pv. tomato

Substrates and Products (Substrate)

EC Number Substrates Comment Substrates Organism Products Comment (Products) Rev. Reac.
2.4.1.10 additional information assay of the growth phenotype of levansucrase-expressing bacteria on agar plate containing sucrose. Only wild-type Lsc3 produces levan giving a mucoid phenotype to the streaks of respective transformant. Quantitative evaluation of the sucrose-splitting (total) levansucrase activity on microplates Pseudomonas syringae pv. tomato ?
-
 ?
2.4.1.10 additional information assay of the growth phenotype of levansucrase-expressing bacteria on agar plate containing sucrose. Only wild-type Lsc3 produces levan giving a mucoid phenotype to the streaks of respective transformant. Quantitative evaluation of the sucrose-splitting (total) levansucrase activity on microplates Pseudomonas syringae pv. tomato DC3000 ?
-
 ?
2.4.1.10 sucrose + alpha-D-glucosyl-(1->2)-[(2->6)-alpha-D-fructosyl]n
-
 Pseudomonas syringae pv. tomato D-glucose + alpha-D-glucosyl-(1->2)-[(2->6)-alpha-D-fructosyl]n+1
-
 ?
2.4.1.10 sucrose + alpha-D-glucosyl-(1->2)-[(2->6)-alpha-D-fructosyl]n
-
 Pseudomonas syringae pv. tomato DC3000 D-glucose + alpha-D-glucosyl-(1->2)-[(2->6)-alpha-D-fructosyl]n+1
-
 ?

Subunits

EC Number Subunits Comment Organism
2.4.1.10 More three-dimensional structure comparison, overview Pseudomonas syringae pv. tomato

Synonyms

EC Number Synonyms Comment Organism
2.4.1.10 Lsc-3
-
 Pseudomonas syringae pv. tomato
2.4.1.10 Lsc3
-
 Pseudomonas syringae pv. tomato

Temperature Optimum [°C]

EC Number Temperature Optimum [°C] Temperature Optimum Maximum [°C] Comment Organism
2.4.1.10 37
-
 assay at Pseudomonas syringae pv. tomato

pH Optimum

EC Number pH Optimum Minimum pH Optimum Maximum Comment Organism
2.4.1.10 6
-
 assay at Pseudomonas syringae pv. tomato

General Information

EC Number General Information Comment Organism
2.4.1.10 metabolism bacterial levansucrases polymerize fructose residues of sucrose to beta-2,6 linked fructans-fructooligosaccharides (FOS) and levan Pseudomonas syringae pv. tomato
2.4.1.10 additional information the enzyme has very high catalytic activity and stability. Key importance of residues Trp109, His113, Glu146 and Glu236 for the catalysis of Lsc3, the catalytic triad is formed by D62, D219, and E303 Pseudomonas syringae pv. tomato