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Literature summary extracted from
- A nitrilase from a metagenomic library acts regioselectively on aliphatic dinitriles (2011), Appl. Microbiol. Biotechnol., 89, 91-98.
Cloned(Commentary)
| EC Number | Cloned (Comment) | Organism |
|---|---|---|
| 3.5.5.1 | gene nit1, DNA and amino acid sequence determination and analysis, expression of His-tagged Nit1 in Escherichia coli strain BL21(DE3) | uncultured bacterium |
| 3.5.5.7 | expressed in Escherichia coli BL21(DE3) cells | uncultured bacterium |
Inhibitors
| EC Number | Inhibitors | Comment | Organism | Structure |
|---|---|---|---|---|
| 3.5.5.7 | Ag+ | 5% residual activity at 1 mM | uncultured bacterium |  |
| 3.5.5.7 | EDTA | 84% residual activity at 1 mM | uncultured bacterium | |
| 3.5.5.7 | Fe2+ | 72% residual activity at 1 mM | uncultured bacterium | |
| 3.5.5.7 | additional information | Nit1 is not or very weakly affected by Na+, Ca2+, Mg2+, and Fe3+ | uncultured bacterium |
KM Value [mM]
| EC Number | KM Value [mM] | KM Value Maximum [mM] | Substrate | Comment | Organism | Structure |
|---|---|---|---|---|---|---|
| 3.5.5.1 | 47 | - |
glutaronitrile | pH 7.0, 30°C, recombinant His-tagged Nit1 | uncultured bacterium | |
| 3.5.5.7 | 47 | - |
glutaronitrile | at 30°C in 50 mM sodium phosphate buffer (pH 7.5) and 1 mM dithiothreitol | uncultured bacterium | |
Molecular Weight [Da]
| EC Number | Molecular Weight [Da] | Molecular Weight Maximum [Da] | Comment | Organism |
|---|---|---|---|---|
| 3.5.5.1 | 38000 | - |
x * 38000, about, SDS-PAGE | uncultured bacterium |
| 3.5.5.7 | 38000 | - |
x * 38000, SDS-PAGE | uncultured bacterium |
| 3.5.5.7 | 700000 | - |
gel filtration | uncultured bacterium |
Organism
| EC Number | Organism | UniProt | Comment | Textmining |
|---|---|---|---|---|
| 3.5.5.1 | uncultured bacterium | E1A0Z9 | gene nit1 | - |
| 3.5.5.7 | uncultured bacterium | E1A0Z9 | - |
- |
Purification (Commentary)
| EC Number | Purification (Comment) | Organism |
|---|---|---|
| 3.5.5.1 | recombinant His-tagged Nit1 from Escherichia coli strain BL21(DE3) by nickel affinity chromatography and gel filtration | uncultured bacterium |
| 3.5.5.7 | Ni-NTA column chromatography and Superdex 200 gel filtration | uncultured bacterium |
Source Tissue
| EC Number | Source Tissue | Comment | Organism | Textmining |
|---|---|---|---|---|
| 3.5.5.1 | additional information | no growth on 4-tolunitrile or 4-hydroxybenzonitrile as sole nitrogen sources | uncultured bacterium | - |
Substrates and Products (Substrate)
| EC Number | Substrates | Comment Substrates | Organism | Products | Comment (Products) | Rev. | Reac. |
|---|---|---|---|---|---|---|---|
| 3.5.5.1 | 2-methylglutaronitrile + 2 H2O | regioselectivity of Nit1 | uncultured bacterium | 4-cyanopentanoic acid + NH3 | - |
? | |
| 3.5.5.1 | 3-cyanopyridine + 2 H2O | - |
uncultured bacterium | pyridine 3-carboxylic acid + NH3 | - |
? | |
| 3.5.5.1 | 3-phenylpropionitrile + H2O | - |
uncultured bacterium | 3-phenylpropionic acid + NH3 | - |
? | |
| 3.5.5.1 | cinnamonitrile + H2O | - |
uncultured bacterium | ? | - |
? | |
| 3.5.5.1 | glutaronitrile + H2O | - |
uncultured bacterium | glutaric acid + NH3 | - |
? | |
| 3.5.5.1 | additional information | Nit1 substrate specificity, overview. Significant activities are observed with the aromatic substrates 3-phenylpropionitrile and cinnamonitrile. To a minor degree, the heterocyclic 3-cyanopyridine is also accepted as substrate. In contrast to aliphatic dinitriles, the aliphatic mononitriles acetonitrile, propionitrile, and butyronitrile are almost not converted by Nit1, no activity on malononitrile | uncultured bacterium | ? | - |
? | |
| 3.5.5.7 | 2-methylglutaronitrile + 2 H2O | 77% activity compared to glutaronitrile | uncultured bacterium | 4-cyanopentanoic acid + NH3 | - |
? | |
| 3.5.5.7 | 3-cyanopyridine + H2O | 11% activity compared to glutaronitrile | uncultured bacterium | 3-carboxypyridine + NH3 | - |
? | |
| 3.5.5.7 | 3-phenylpropionitrile + H2O | 59% activity compared to glutaronitrile | uncultured bacterium | ? | - |
? | |
| 3.5.5.7 | acetonitrile + H2O | 5% activity compared to glutaronitrile | uncultured bacterium | ? | - |
? | |
| 3.5.5.7 | adiponitrile + H2O | 91% activity compared to glutaronitrile | uncultured bacterium | adipate + NH3 | - |
? | |
| 3.5.5.7 | butyronitrile + H2O | 6% activity compared to glutaronitrile | uncultured bacterium | ? | - |
? | |
| 3.5.5.7 | cinnamonitrile + H2O | 28% activity compared to glutaronitrile | uncultured bacterium | cinnamate + NH3 | - |
? | |
| 3.5.5.7 | crotonitrile + H2O | 7% activity compared to glutaronitrile | uncultured bacterium | ? | - |
? | |
| 3.5.5.7 | glutaronitrile + H2O | 100% activity | uncultured bacterium | glutarate + NH3 | - |
? | |
| 3.5.5.7 | additional information | Nit1 does not hydrolyze malononitrile, mandelonitrile, 4-cyanobenzoic acid, 3-ethoxybenzonitrile, 4-hydroxybenzonitrile, alpha-methylbenzyl cyanide, acrylonitrile, 4-tolunitrile, and benzonitrile | uncultured bacterium | ? | - |
? | |
| 3.5.5.7 | additional information | Nit1 is an aliphatic nitrilase favoring dinitriles over mononitriles | uncultured bacterium | ? | - |
? | |
| 3.5.5.7 | propionitrile + H2O | 5% activity compared to glutaronitrile | uncultured bacterium | ? | - |
? | |
| 3.5.5.7 | succinonitrile + H2O | 76% activity compared to glutaronitrile | uncultured bacterium | succinate + NH3 | - |
? | |
Subunits
| EC Number | Subunits | Comment | Organism |
|---|---|---|---|
| 3.5.5.1 | ? | x * 38000, about, SDS-PAGE | uncultured bacterium |
| 3.5.5.7 | ? | x * 38000, SDS-PAGE | uncultured bacterium |
Synonyms
| EC Number | Synonyms | Comment | Organism |
|---|---|---|---|
| 3.5.5.1 | NIT1 | - |
uncultured bacterium |
| 3.5.5.1 | nitrilase | - |
uncultured bacterium |
| 3.5.5.7 | NIT1 | - |
uncultured bacterium |
Temperature Optimum [°C]
| EC Number | Temperature Optimum [°C] | Temperature Optimum Maximum [°C] | Comment | Organism |
|---|---|---|---|---|
| 3.5.5.1 | 30 | - |
recombinant His-tagged Nit1 | uncultured bacterium |
| 3.5.5.7 | 30 | - |
using the substrate glutaronitrile | uncultured bacterium |
Temperature Stability [°C]
| EC Number | Temperature Stability Minimum [°C] | Temperature Stability Maximum [°C] | Comment | Organism |
|---|---|---|---|---|
| 3.5.5.1 | 40 | - |
purified recombinant His-tagged Nit1, 1 h, stable | uncultured bacterium |
| 3.5.5.1 | 50 | - |
purified recombinant His-tagged Nit1, 30 min, loss of 50% activity | uncultured bacterium |
| 3.5.5.1 | 60 | - |
purified recombinant His-tagged Nit1, 10 in, complete inactivation | uncultured bacterium |
| 3.5.5.7 | 40 | 60 | up to 40°C Nit1 is stable for 1 h, while the loss of activity is about 50% after 30 min at 50°C, the enzyme is completely inactivated after 10 min incubation at 60°C | uncultured bacterium |
pH Optimum
| EC Number | pH Optimum Minimum | pH Optimum Maximum | Comment | Organism |
|---|---|---|---|---|
| 3.5.5.1 | 7 | - |
recombinant His-tagged Nit1 | uncultured bacterium |
| 3.5.5.7 | 7 | - |
using the substrate glutaronitrile | uncultured bacterium |
pH Range
| EC Number | pH Minimum | pH Maximum | Comment | Organism |
|---|---|---|---|---|
| 3.5.5.1 | 5 | 9 | over 50% of maximal activity within this range | uncultured bacterium |
| 3.5.5.7 | 5 | 9 | Nit1 tolerates a pH range between 5 and 9 with an activity of more than 50% | uncultured bacterium |
kcat/KM [mM/s]
| EC Number | kcat/KM Value [1/mMs-1] | kcat/KM Value Maximum [1/mMs-1] | Substrate | Comment | Organism | Structure |
|---|---|---|---|---|---|---|
| 3.5.5.1 | 0.48 | - |
glutaronitrile | pH 7.0, 30°C, recombinant His-tagged Nit1 | uncultured bacterium | |
| 3.5.5.7 | 0.483 | - |
glutaronitrile | at 30°C in 50 mM sodium phosphate buffer (pH 7.5) and 1 mM dithiothreitol | uncultured bacterium | |
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