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Literature summary for 4.1.1.15 extracted from
- Pyridoxine supplementation improves the activity of recombinant glutamate decarboxylase and the enzymatic production of gama-aminobutyric acid (2016), PLoS ONE, 11, e0157466 .
Cloned(Commentary)
| Cloned (Comment) | Organism |
|---|---|
| gene gadB, recombinant cytoplasmic expression in Escherichia coli strain BL21(DE3), using vectors pMD18-T and pET-24a(+) and IPTG induction, subcloning in Escherichia coli strain JM109 | Escherichia coli |
Protein Variants
| Protein Variants | Comment | Organism |
|---|---|---|
| additional information | optimization of the reaction conditions for recombinant GABA production by the homogenously expressed enzyme from Escherichia coli. The activity for GAD produced in media supplemented with 0.05 mM soluble vitamin B6 analogue pyridoxine hydrochloride (GAD-V) is 154.8 U/l, 1.8fold higher than that of GAD obtained without supplementation (GAD-C). Purified GAD-V exhibits increased activity of 193.4 U/mg (1.5fold higher), superior thermostability (2.8fold greater), and higher kcat/Km (1.6fold higher) compared to GAD-C. Under optimal conditions in reactions mixtures lacking added pyridoxal 5'-phosphate, crude GAD-V converts 500 g/l monosodium glutamate to GABA with a yield of 100%, and 750 g/l monosodium glutamate with a yield of 88.7%. Effect of substrate concentration on GABA production by GAD-C and GAD-V, overview | Escherichia coli |
Localization
| Localization | Comment | Organism | GeneOntology No. | Textmining |
|---|---|---|---|---|
| cytoplasm | - |
Escherichia coli | 5737 | - |
Molecular Weight [Da]
| Molecular Weight [Da] | Molecular Weight Maximum [Da] | Comment | Organism |
|---|---|---|---|
| 324000 | - |
recombinant enzyme, gel filtration | Escherichia coli |
Natural Substrates/ Products (Substrates)
| Natural Substrates | Organism | Comment (Nat. Sub.) | Natural Products | Comment (Nat. Pro.) | Rev. | Reac. |
|---|---|---|---|---|---|---|
| L-glutamate | Escherichia coli | - |
4-aminobutanoate + CO2 | - |
ir |  |
| L-glutamate | Escherichia coli K-12 / MG1655 | - |
4-aminobutanoate + CO2 | - |
ir | |
Organism
| Organism | UniProt | Comment | Textmining |
|---|---|---|---|
| Escherichia coli | P69910 | - |
- |
| Escherichia coli K-12 / MG1655 | P69910 | - |
- |
Purification (Commentary)
| Purification (Comment) | Organism |
|---|---|
| recombinant enzyme 1.7-1.9fold from Escherichia coli strain BL21(DE3) by ammonium sulfate fractionation, dialysis, anion exchange chromatographyand again dialysis, to homogeneity | Escherichia coli |
Specific Activity [micromol/min/mg]
| Specific Activity Minimum [µmol/min/mg] | Specific Activity Maximum [µmol/min/mg] | Comment | Organism |
|---|---|---|---|
| 130.2 | - |
purified recombinant GAD obtained without supplementation (GAD-C), pH 4.8, 30°C | Escherichia coli |
| 193.4 | - |
purified recombinant GAD produced in media supplemented with 0.05 mM soluble vitamin B6 analogue pyridoxine hydrochloride (GAD-V), pH 4.8, 30°C | Escherichia coli |
Substrates and Products (Substrate)
| Substrates | Comment Substrates | Organism | Products | Comment (Products) | Rev. | Reac. |
|---|---|---|---|---|---|---|
| L-glutamate | - |
Escherichia coli | 4-aminobutanoate + CO2 | - |
ir | |
| L-glutamate | - |
Escherichia coli K-12 / MG1655 | 4-aminobutanoate + CO2 | - |
ir | |
Subunits
| Subunits | Comment | Organism |
|---|---|---|
| hexamer | 6 * 54000, recombinant enzyme, SDS-PAGE | Escherichia coli |
| More | addition of pyridoxine does not influence the aggregation state of GAD | Escherichia coli |
Synonyms
| Synonyms | Comment | Organism |
|---|---|---|
| GAD | - |
Escherichia coli |
| GadB | - |
Escherichia coli |
Temperature Optimum [°C]
| Temperature Optimum [°C] | Temperature Optimum Maximum [°C] | Comment | Organism |
|---|---|---|---|
| 50 | - |
- |
Escherichia coli |
Temperature Range [°C]
| Temperature Minimum [°C] | Temperature Maximum [°C] | Comment | Organism |
|---|---|---|---|
| 30 | 60 | activity range | Escherichia coli |
Temperature Stability [°C]
| Temperature Stability Minimum [°C] | Temperature Stability Maximum [°C] | Comment | Organism |
|---|---|---|---|
| 50 | - |
GAD-C and GAD-V, stable up to, rapid loss of activity above | Escherichia coli |
pH Optimum
| pH Optimum Minimum | pH Optimum Maximum | Comment | Organism |
|---|---|---|---|
| 5 | - |
- |
Escherichia coli |
pH Range
| pH Minimum | pH Maximum | Comment | Organism |
|---|---|---|---|
| 3 | 7 | purified recombinant enzyme, the activity decreases dramatically between pH 5.0 and pH 7.0. GAD-V exhibits a much higher activity than GAD-C below pH 5.0. More than 80% of the maximum catalytic activity is observed for GAD-V between pH 4.0 and pH 5.0, while GAD-C exhibits 98.3% and 38% of the activity shown by GAD-V at pH 5.0 and 4.0, respectively | Escherichia coli |
pH Stability
| pH Stability | pH Stability Maximum | Comment | Organism |
|---|---|---|---|
| 3 | 7 | recombinant enzyme, 4°C, 24 h, both GAD-C and GAD-V retain more than 70% of their maximal activity at pH 3.0-5.0 and more than 80% at pH 5.0-7.0 | Escherichia coli |
Cofactor
| Cofactor | Comment | Organism | Structure |
|---|---|---|---|
| pyridoxal 5'-phosphate | - |
Escherichia coli | |
General Information
| General Information | Comment | Organism |
|---|---|---|
| physiological function | glutamate decarboxylase (GAD) catalyzes the irreversible decarboxylation of L-glutamate to the valuable food supplement gamma-aminobutyric acid (GABA) | Escherichia coli |
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