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Literature summary for 1.14.14.10 extracted from
- Purification and characterization of a two-component monooxygenase that hydroxylates nitrilotriacetate from "Chelatobacter" strain ATCC 29600 (1992), J. Bacteriol., 174, 1179-1188.
Activating Compound
| Activating Compound | Comment | Organism | Structure |
|---|---|---|---|
| component A of NTA monooxygenase | the ability of component B of NTA monooxygenase to catalyze the oxidation of nitrilotriacetate to iminodiacetate and glyoxylate is completely dependent on the presence of component A | Aminobacter aminovorans | |
| FMN | NTA-Mo activity increases by a factor of 4 when 0.003 mM FMN is included in the assay mixture, there is 0.4 mol of FMN per mol of purified component B | Aminobacter aminovorans |  |
| additional information | the activity is increased neither by additions of 20% (w/v) polyethylene glycol or sorbitol | Aminobacter aminovorans |
Inhibitors
| Inhibitors | Comment | Organism | Structure |
|---|---|---|---|
| 5,5'-dithiobis(2-nitrobenzoate) | 67% residual activity at 1 mM | Aminobacter aminovorans | |
| Ca2+ | 5% residual activity at 10 mM | Aminobacter aminovorans | |
| EDTA | 3% residual activity at 10 mM | Aminobacter aminovorans | |
| FAD | 24% residual activity at 0.1 mM | Aminobacter aminovorans | |
| NaCl | 54% residual activity at 250 mM | Aminobacter aminovorans | |
| p-hydroxymercuribenzoate | 12% residual activity at 1 mM | Aminobacter aminovorans | |
KM Value [mM]
| KM Value [mM] | KM Value Maximum [mM] | Substrate | Comment | Organism | Structure |
|---|---|---|---|---|---|
| 0.0013 | - |
FMN | in 40 mM TAPS buffer at pH 8.5 and 25°C | Aminobacter aminovorans | |
| 0.5 | - |
nitrilotriacetate | in 40 mM TAPS buffer at pH 8.5 and 25°C | Aminobacter aminovorans | |
Metals/Ions
| Metals/Ions | Comment | Organism | Structure |
|---|---|---|---|
| Co2+ | may replace Mg2+, enzyme activity is reduced to 80% of that with MgCl2 | Aminobacter aminovorans | |
| Mg2+ | required, 2 mM used in assay conditions | Aminobacter aminovorans | |
| Mn2+ | Mn2+ ions are able to replace Mg2+ but lead to a higher uncoupled NADH oxidation and enzyme activity is reduced to 70% of that with MgCl2 | Aminobacter aminovorans | |
| additional information | no nitrilotriacetate consumption is observed with Ca2+,Fe2+, Fe3+, Zn2+, Cu2+, or Ni2+ | Aminobacter aminovorans | |
| additional information | the enzyme contains less than 0.15 atom of Fe per mol of protein, indicating that neither Fe-sulfur clusters nor cytochromes are present | Aminobacter aminovorans |
Molecular Weight [Da]
| Molecular Weight [Da] | Molecular Weight Maximum [Da] | Comment | Organism |
|---|---|---|---|
| 36000 | - |
2 * 36000, component B of NTA monooxygenase, SDS-PAGE | Aminobacter aminovorans |
Organism
| Organism | UniProt | Comment | Textmining |
|---|---|---|---|
| Aminobacter aminovorans | - |
- |
- |
| Aminobacter aminovorans ATCC 29600 | - |
- |
- |
Purification (Commentary)
| Purification (Comment) | Organism |
|---|---|
| ammonium sulfate precipitation, phenyl Sepharose column chromatography, TMAE-Fractogel column chromatography, and phenyl Superose column chromatography | Aminobacter aminovorans |
Storage Stability
| Storage Stability | Organism |
|---|---|
| -70°C, purified component B of NTA monooxygenase in 20 mM HEPES buffer with 2 mM dithiothreitol, at pH 7.8, several months, without any significant loss of activity | Aminobacter aminovorans |
| 4°C, crude extract, 50 mM Tris-HCl, 200 h, 50% loss of activity | Aminobacter aminovorans |
| 4°C, purified component B of NTA monooxygenase in 20 mM HEPES buffer at pH 7.8, 7 days, 70% loss of activity | Aminobacter aminovorans |
| 4°C, purified component B of NTA monooxygenase in 20 mM HEPES buffer with 2 mM dithiothreitol and ammonium sulfate to 5% saturation, at pH 7.8, 7 days, 5% loss of activity | Aminobacter aminovorans |
| 4°C, purified component B of NTA monooxygenase in 20 mM HEPES buffer with 2 mM dithiothreitol, at pH 7.8, 7 days, 20% loss of activity | Aminobacter aminovorans |
Substrates and Products (Substrate)
| Substrates | Comment Substrates | Organism | Products | Comment (Products) | Rev. | Reac. |
|---|---|---|---|---|---|---|
| additional information | less than 2% reactivity with N-methyl-iminodiacetate, N-acetamido-iminodiacetate, N-hydroxyethyl-iminodiacetate, EDTA, trimethylamine, triethanolamine, dimethylamine, dimethylglycine, methylamine, sarcosine, anthranilate, betaine, N-methylglutamate, N-methylalanine, N-methylaspartate, N-acetylglutamate, N-methylphenylalanine, N-methylleucine, N-methyltryptophane, glutamate, aspartate, citrate, proline, imidazoleacetate, asparagine, and ethylenediamine-N,N'-diacetate | Aminobacter aminovorans | ? | - |
? | |
| additional information | less than 2% reactivity with N-methyl-iminodiacetate, N-acetamido-iminodiacetate, N-hydroxyethyl-iminodiacetate, EDTA, trimethylamine, triethanolamine, dimethylamine, dimethylglycine, methylamine, sarcosine, anthranilate, betaine, N-methylglutamate, N-methylalanine, N-methylaspartate, N-acetylglutamate, N-methylphenylalanine, N-methylleucine, N-methyltryptophane, glutamate, aspartate, citrate, proline, imidazoleacetate, asparagine, and ethylenediamine-N,N'-diacetate | Aminobacter aminovorans ATCC 29600 | ? | - |
? | |
| nitrilotriacetate + FMNH2 + H+ + O2 | specific substrate | Aminobacter aminovorans | iminodiacetate + glyoxylate + FMN + H2O | - |
? | |
| nitrilotriacetate + FMNH2 + H+ + O2 | specific substrate | Aminobacter aminovorans ATCC 29600 | iminodiacetate + glyoxylate + FMN + H2O | - |
? | |
Subunits
| Subunits | Comment | Organism |
|---|---|---|
| homodimer | 2 * 36000, component B of NTA monooxygenase, SDS-PAGE | Aminobacter aminovorans |
Synonyms
| Synonyms | Comment | Organism |
|---|---|---|
| component B of NTA monooxygenase | - |
Aminobacter aminovorans |
| NTA monooxygenase | - |
Aminobacter aminovorans |
| NTA-MO | - |
Aminobacter aminovorans |
Temperature Optimum [°C]
| Temperature Optimum [°C] | Temperature Optimum Maximum [°C] | Comment | Organism |
|---|---|---|---|
| 25 | - |
12% activity is left at 2°C, and no activity is observed at 40°C | Aminobacter aminovorans |
pH Optimum
| pH Optimum Minimum | pH Optimum Maximum | Comment | Organism |
|---|---|---|---|
| 7.8 | - |
pH optimum for NTA consumption in cell extracts. At pH 7.2 and 8.4, 25% of the maximum activity is obtained, and no activity is left at pH 6.0 or 9.0 | Aminobacter aminovorans |
| 8.5 | - |
pH optimum for the purified enzyme | Aminobacter aminovorans |
pH Range
| pH Minimum | pH Maximum | Comment | Organism |
|---|---|---|---|
| 7 | 9.6 | 35% activity of the purified enzyme remains at pH 7.0 and 9.6 | Aminobacter aminovorans |
Cofactor
| Cofactor | Comment | Organism | Structure |
|---|---|---|---|
| FMN | omission of FMN or replacement of FMN by FAD results in a residual activity of 2% when component B is not resaturated with FMN | Aminobacter aminovorans | |
| NADH | NADPH cannot replace NADH | Aminobacter aminovorans | |
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