Any feedback?
Literature summary extracted from
- The structural basis of Erwinia rhapontici isomaltulose synthase (2013), PLoS ONE, 8, e74788.
Cloned(Commentary)
| EC Number | Cloned (Comment) | Organism |
|---|---|---|
| 5.4.99.11 | expression of His-tagged wild-type and mutant enzymes in Escherichia coli strain BL21(DE3) | Erwinia rhapontici |
Crystallization (Commentary)
| EC Number | Crystallization (Comment) | Organism |
|---|---|---|
| 5.4.99.11 | wild-type NX-5 and enzyme mutant complexes E295Q-sucrose and D241A-glucose, sitting drop vapor diffusion method, mixing of 0.001 ml of 10 mg/ml protein solution with 0.001 ml of reservoir solution containing 0.1 M trisodium citrate, pH 5.6, 0.2 M ammonium acetate, and 30% PEG 4000, and for sucrose-complexed mutant enzyme crystal 20 mM sucrose, 0.1 M bicine, pH 9.0, and 30% PEG 6000. The D241A-glucose complex and R335H/R336T/K337I/D338P-glucose complex are co-crystallized in the presence of 20 mM sucrose, 0.1 M sodium cacodylate, pH 6.5, and 29% PEG 8000, 20°C, 3-5 days, X-ray diffraction structure determination and analysis at 1.70 A, 1.70 a, and 2.00 A resolutions, respectively | Erwinia rhapontici |
Protein Variants
| EC Number | Protein Variants | Comment | Organism |
|---|---|---|---|
| 5.4.99.11 | D241A | site-directed mutagenesis, glucose binding structure in comparison to the wild-type enzyme by crystal structure analysis | Erwinia rhapontici |
| 5.4.99.11 | E295Q | site-directed mutagenesis, sucrose binding structure in comparison to the wild-type enzyme by crystal structure analysis | Erwinia rhapontici |
| 5.4.99.11 | F297A | site-directed mutagenesis, the mutant shows an altered product ratio of trehalulose and isomaltulose compared to the wild-type enzyme | Erwinia rhapontici |
| 5.4.99.11 | F321A | site-directed mutagenesis, the mutant shows an altered product ratio of trehalulose and isomaltulose compared to the wild-type enzyme | Erwinia rhapontici |
| 5.4.99.11 | additional information | mutations of the loop region of enzyme NX-5 result in significant changes of the product ratio between isomaltulose and trehalulose | Erwinia rhapontici |
| 5.4.99.11 | R325D | site-directed mutagenesis, the mutant shows an altered product ratio of trehalulose and isomaltulose compared to the wild-type enzyme | Erwinia rhapontici |
| 5.4.99.11 | R328D | site-directed mutagenesis, the mutant shows an altered product ratio of trehalulose and isomaltulose compared to the wild-type enzyme | Erwinia rhapontici |
| 5.4.99.11 | R335H/R336T/K337I/D338P | site-directed mutagenesis, glucose binding structure in comparison to the wild-type enzyme by crystal structure analysis | Erwinia rhapontici |
Natural Substrates/ Products (Substrates)
| EC Number | Natural Substrates | Organism | Comment (Nat. Sub.) | Natural Products | Comment (Nat. Pro.) | Rev. | Reac. |
|---|---|---|---|---|---|---|---|
| 5.4.99.11 | additional information | Erwinia rhapontici | the enzyme catalyzes the isomerization of sucrose (alpha-D-glucosylpyranosyl-1,2-beta-D-fructofuranose) to isomaltulose (alpha-D-glucosylpyranosyl-1,6-D-fructofuranose) as the main product, and trehalulose (alpha-D-glucosylpyranosyl-1,1-D-fructofuranose) as minor product, and produces glucose and fructose in residual amounts because of sucrose hydrolysis | ? | - |
? |  |
| 5.4.99.11 | additional information | Erwinia rhapontici NX-5 | the enzyme catalyzes the isomerization of sucrose (alpha-D-glucosylpyranosyl-1,2-beta-D-fructofuranose) to isomaltulose (alpha-D-glucosylpyranosyl-1,6-D-fructofuranose) as the main product, and trehalulose (alpha-D-glucosylpyranosyl-1,1-D-fructofuranose) as minor product, and produces glucose and fructose in residual amounts because of sucrose hydrolysis | ? | - |
? | |
| 5.4.99.11 | sucrose | Erwinia rhapontici | product specificity of enzyme NX-5 towards isomaltulose and the role of the loop330-339 in NX-5 catalysis | 1-O-alpha-D-glucopyranosyl-D-fructofuranose + alpha-D-glucosylpyranosyl-1,6-D-fructofuranose | - |
? | |
| 5.4.99.11 | sucrose | Erwinia rhapontici NX-5 | product specificity of enzyme NX-5 towards isomaltulose and the role of the loop330-339 in NX-5 catalysis | 1-O-alpha-D-glucopyranosyl-D-fructofuranose + alpha-D-glucosylpyranosyl-1,6-D-fructofuranose | - |
? | |
Organism
| EC Number | Organism | UniProt | Comment | Textmining |
|---|---|---|---|---|
| 5.4.99.11 | Erwinia rhapontici | - |
- |
- |
| 5.4.99.11 | Erwinia rhapontici NX-5 | - |
- |
- |
Purification (Commentary)
| EC Number | Purification (Comment) | Organism |
|---|---|---|
| 5.4.99.11 | recombinant His-tagged wild-type and mutant enzymes from Escherichia coli strain BL21(DE3) by nickel affinity chromatograhy and gel filtration | Erwinia rhapontici |
Substrates and Products (Substrate)
| EC Number | Substrates | Comment Substrates | Organism | Products | Comment (Products) | Rev. | Reac. |
|---|---|---|---|---|---|---|---|
| 5.4.99.11 | additional information | the enzyme catalyzes the isomerization of sucrose (alpha-D-glucosylpyranosyl-1,2-beta-D-fructofuranose) to isomaltulose (alpha-D-glucosylpyranosyl-1,6-D-fructofuranose) as the main product, and trehalulose (alpha-D-glucosylpyranosyl-1,1-D-fructofuranose) as minor product, and produces glucose and fructose in residual amounts because of sucrose hydrolysis | Erwinia rhapontici | ? | - |
? | |
| 5.4.99.11 | additional information | the enzyme catalyzes the isomerization of sucrose (alpha-D-glucosylpyranosyl-1,2-beta-D-fructofuranose) to isomaltulose (alpha-D-glucosylpyranosyl-1,6-D-fructofuranose) as the main product, and trehalulose (alpha-D-glucosylpyranosyl-1,1-D-fructofuranose) as minor product, and produces glucose and fructose in residual amounts because of sucrose hydrolysis | Erwinia rhapontici NX-5 | ? | - |
? | |
| 5.4.99.11 | sucrose | product specificity of enzyme NX-5 towards isomaltulose and the role of the loop330-339 in NX-5 catalysis | Erwinia rhapontici | 1-O-alpha-D-glucopyranosyl-D-fructofuranose + alpha-D-glucosylpyranosyl-1,6-D-fructofuranose | - |
? | |
| 5.4.99.11 | sucrose | product specificity of enzyme NX-5 towards isomaltulose and the role of the loop330-339 in NX-5 catalysis | Erwinia rhapontici | 1-O-alpha-D-glucopyranosyl-D-fructofuranose + alpha-D-glucosylpyranosyl-1,6-D-fructofuranose | main product isomaltulose, i.e. alpha-D-glucopyranosyl-1,6-D-fructofuranose or palatinose, and by-product trehalulose, i.e. alpha-D-glucopyranosyl-1,1-D-fructofuranose are two structural isomers of sucrose | ? | |
| 5.4.99.11 | sucrose | product specificity of enzyme NX-5 towards isomaltulose and the role of the loop330-339 in NX-5 catalysis | Erwinia rhapontici NX-5 | 1-O-alpha-D-glucopyranosyl-D-fructofuranose + alpha-D-glucosylpyranosyl-1,6-D-fructofuranose | - |
? | |
| 5.4.99.11 | sucrose | product specificity of enzyme NX-5 towards isomaltulose and the role of the loop330-339 in NX-5 catalysis | Erwinia rhapontici NX-5 | 1-O-alpha-D-glucopyranosyl-D-fructofuranose + alpha-D-glucosylpyranosyl-1,6-D-fructofuranose | main product isomaltulose, i.e. alpha-D-glucopyranosyl-1,6-D-fructofuranose or palatinose, and by-product trehalulose, i.e. alpha-D-glucopyranosyl-1,1-D-fructofuranose are two structural isomers of sucrose | ? | |
Subunits
| EC Number | Subunits | Comment | Organism |
|---|---|---|---|
| 5.4.99.11 | More | enzyme structural analysis, overview | Erwinia rhapontici |
Synonyms
| EC Number | Synonyms | Comment | Organism |
|---|---|---|---|
| 5.4.99.11 | NX-5 | - |
Erwinia rhapontici |
| 5.4.99.11 | SIase | - |
Erwinia rhapontici |
| 5.4.99.11 | sucrose isomerase | - |
Erwinia rhapontici |
Temperature Optimum [°C]
| EC Number | Temperature Optimum [°C] | Temperature Optimum Maximum [°C] | Comment | Organism |
|---|---|---|---|---|
| 5.4.99.11 | 30 | - |
assay at | Erwinia rhapontici |
pH Optimum
| EC Number | pH Optimum Minimum | pH Optimum Maximum | Comment | Organism |
|---|---|---|---|---|
| 5.4.99.11 | 6 | - |
assay at | Erwinia rhapontici |
General Information
| EC Number | General Information | Comment | Organism |
|---|---|---|---|
| 5.4.99.11 | additional information | structural basis for the product specificity of NX-5, docking of fructofuranose and fructopyranose into the active site of the enzyme mutant D241A-glucose complex, molecular dynamics simulations, molecular mechanism controlling sucrose isomer formation, overview. Phe297 and Phe321 in enzyme NX-5 form an aromatic clamp near the entrance of the active pocket. Residues Asp102, His145, Arg239, His368, Asp369, Glu428, and Arg456 are responsible for sucrose binding and residues Asp241 (the nucleophile) and Glu295 (the acid catalyst) are crucial for sucrose hydrolysis. The active site pocket is stabilized by the salt bridge interactions between Arg239-Asp241, Asp102-Arg456 and Asp369-Arg456. Residues Arg325 and Arg328 are located in the325RLDRD329 motif and play roles in the product specificity | Erwinia rhapontici |
Use of this online version of BRENDA is free under the CC BY 4.0 license. See terms of use for full details.
Release 2023.1 (January 2023)
Legal
Curated at
Member of
