Cloned (Comment) | Organism |
---|---|
gene tnaA, recombinant expression of His6-tagged enzyme VcTrpase in Escherichia coli strain BL21(DE3) tn5:tnaA | Vibrio cholerae O1 |
Protein Variants | Comment | Organism |
---|---|---|
D137A | site-directed mutagenesis, inactive mutant | Vibrio cholerae O1 |
H463A | site-directed mutagenesis, the mutant shows 96% reduced activity compared to the wild-type | Vibrio cholerae O1 |
K269A | site-directed mutagenesis, inactive mutant | Vibrio cholerae O1 |
K270A | site-directed mutagenesis, the mutation possibly eliminates this cofactor-protein interaction in the enzyme, inactive mutant | Vibrio cholerae O1 |
R103A | site-directed mutagenesis, the mutation alters orientation of the cofactor pyridoxal 5'-phosphate, inactive mutant | Vibrio cholerae O1 |
R230A | site-directed mutagenesis, inactive mutant | Vibrio cholerae O1 |
Y52A | site-directed mutagenesis, inactive mutant | Vibrio cholerae O1 |
Y74A | site-directed mutagenesis, inactive mutant | Vibrio cholerae O1 |
KM Value [mM] | KM Value Maximum [mM] | Substrate | Comment | Organism | Structure |
---|---|---|---|---|---|
additional information | - |
additional information | Michaelis-Menten kinetics | Vibrio cholerae O1 | |
0.346 | - |
S-Benzyl-L-cysteine | recombinant His-tagged wild-type enzyme, pH 7.8, 37°C | Vibrio cholerae O1 | |
0.49 | - |
S-methyl-L-cysteine | recombinant His-tagged wild-type enzyme, pH 7.8, 37°C | Vibrio cholerae O1 | |
5.252 | - |
L-tryptophan | recombinant His-tagged wild-type enzyme, pH 7.8, 37°C | Vibrio cholerae O1 |
Natural Substrates | Organism | Comment (Nat. Sub.) | Natural Products | Comment (Nat. Pro.) | Rev. | Reac. |
---|---|---|---|---|---|---|
L-tryptophan + H2O | Vibrio cholerae O1 | - |
indole + pyruvate + NH3 | - |
? | |
L-tryptophan + H2O | Vibrio cholerae O1 ATCC 39541 | - |
indole + pyruvate + NH3 | - |
? |
Organism | UniProt | Comment | Textmining |
---|---|---|---|
Vibrio cholerae O1 | A5EYI0 | - |
- |
Vibrio cholerae O1 ATCC 39541 | A5EYI0 | - |
- |
Purification (Comment) | Organism |
---|---|
recombinant His6-tagged enzyme VcTrpase from Escherichia coli strain BL21(DE3) tn5:tnaA by nickel affinity chromatography | Vibrio cholerae O1 |
Substrates | Comment Substrates | Organism | Products | Comment (Products) | Rev. | Reac. |
---|---|---|---|---|---|---|
L-tryptophan + H2O | - |
Vibrio cholerae O1 | indole + pyruvate + NH3 | - |
? | |
L-tryptophan + H2O | - |
Vibrio cholerae O1 ATCC 39541 | indole + pyruvate + NH3 | - |
? | |
additional information | coupled assay: the decrease in NADH absorbance at 340 nm in the presence of LDH is used to determine Trpase activity after adding the substrate. L-Phenylalanine and L-serine are no substrates | Vibrio cholerae O1 | ? | - |
? | |
additional information | coupled assay: the decrease in NADH absorbance at 340 nm in the presence of LDH is used to determine Trpase activity after adding the substrate. L-Phenylalanine and L-serine are no substrates | Vibrio cholerae O1 ATCC 39541 | ? | - |
? | |
S-benzyl-L-cysteine + H2O | - |
Vibrio cholerae O1 | phenylmethanethiol + pyruvate + NH3 | - |
? | |
S-benzyl-L-cysteine + H2O | - |
Vibrio cholerae O1 ATCC 39541 | phenylmethanethiol + pyruvate + NH3 | - |
? | |
S-methyl-L-cysteine + H2O | - |
Vibrio cholerae O1 | methanethiol + pyruvate + NH3 | - |
? | |
S-methyl-L-cysteine + H2O | - |
Vibrio cholerae O1 ATCC 39541 | methanethiol + pyruvate + NH3 | - |
? |
Subunits | Comment | Organism |
---|---|---|
? | x * 49000, His6-tagged enzyme VcTrpase, SDS-PAGE | Vibrio cholerae O1 |
Synonyms | Comment | Organism |
---|---|---|
TnaA | - |
Vibrio cholerae O1 |
Trpase | - |
Vibrio cholerae O1 |
VcTrpase | - |
Vibrio cholerae O1 |
Temperature Optimum [°C] | Temperature Optimum Maximum [°C] | Comment | Organism |
---|---|---|---|
45 | - |
- |
Vibrio cholerae O1 |
Temperature Minimum [°C] | Temperature Maximum [°C] | Comment | Organism |
---|---|---|---|
22 | 70 | activity range, inactivation above | Vibrio cholerae O1 |
Temperature Stability Minimum [°C] | Temperature Stability Maximum [°C] | Comment | Organism |
---|---|---|---|
4 | - |
VcTrpase does not dissociate into inactive dimers or monomers at 4°C | Vibrio cholerae O1 |
Turnover Number Minimum [1/s] | Turnover Number Maximum [1/s] | Substrate | Comment | Organism | Structure |
---|---|---|---|---|---|
0.346 | - |
S-Benzyl-L-cysteine | recombinant His-tagged wild-type enzyme, pH 7.8, 37°C | Vibrio cholerae O1 | |
0.49 | - |
S-methyl-L-cysteine | recombinant His-tagged wild-type enzyme, pH 7.8, 37°C | Vibrio cholerae O1 | |
5.252 | - |
L-tryptophan | recombinant His-tagged wild-type enzyme, pH 7.8, 37°C | Vibrio cholerae O1 |
pH Optimum Minimum | pH Optimum Maximum | Comment | Organism |
---|---|---|---|
9 | - |
- |
Vibrio cholerae O1 |
pH Minimum | pH Maximum | Comment | Organism |
---|---|---|---|
6 | 9 | high activity within this range | Vibrio cholerae O1 |
pH Stability | pH Stability Maximum | Comment | Organism |
---|---|---|---|
6.5 | 8 | purified recombinant His-tagged enzyme, 10 min, over 80% activity remaining | Vibrio cholerae O1 |
Cofactor | Comment | Organism | Structure |
---|---|---|---|
pyridoxal 5'-phosphate | dependent on, residue Arg103 plays an important role in orientation of the cofactor pyridoxal 5'-phosphate | Vibrio cholerae O1 |
General Information | Comment | Organism |
---|---|---|
additional information | eight catalytically important residues Thr52, Tyr74, Arg103, Asp137, Arg230, Lys269, Lys270, and His463 are located close to the Trpase active site and are absolutely conserved in Trpases. Five of them are located in the conserved regions and are reported to confer a crucial role for binding of the substrate and cofactor to produce the formation of the best intermediate that will lead to substrate degradation. Despite the apparent diversity in the protein sequences, these regions may be essential for enzyme activity to generate indole, which is an important agent for bacterial physiology, ecological balance, and virulence | Vibrio cholerae O1 |
physiological function | the enzyme is responsible for the production of indole, an important intra- and interspecies signaling molecule in bacteria | Vibrio cholerae O1 |
kcat/KM Value [1/mMs-1] | kcat/KM Value Maximum [1/mMs-1] | Substrate | Comment | Organism | Structure |
---|---|---|---|---|---|
0.016 | - |
S-methyl-L-cysteine | recombinant His-tagged wild-type enzyme, pH 7.8, 37°C | Vibrio cholerae O1 | |
1.648 | - |
S-Benzyl-L-cysteine | recombinant His-tagged wild-type enzyme, pH 7.8, 37°C | Vibrio cholerae O1 | |
8.582 | - |
L-tryptophan | recombinant His-tagged wild-type enzyme, pH 7.8, 37°C | Vibrio cholerae O1 |