Cloned (Comment) | Organism |
---|---|
gene Vmut_0738, DNA and amino acid sequence determination and analysis, sequence comparisons, recombinant expression of His6-tagged enzyme in Escherichia coli strain DLT1270 from vector pQE30_Vmut0738 | Vulcanisaeta moutnovskia |
KM Value [mM] | KM Value Maximum [mM] | Substrate | Comment | Organism | Structure |
---|---|---|---|---|---|
0.84 | - |
L-norvaline | pH 8.0, 65°C | Vulcanisaeta moutnovskia | |
1.17 | - |
L-valine | pH 8.0, 65°C | Vulcanisaeta moutnovskia | |
7.95 | - |
L-ornithine | pH 8.0, 65°C | Vulcanisaeta moutnovskia |
Molecular Weight [Da] | Molecular Weight Maximum [Da] | Comment | Organism |
---|---|---|---|
145000 | - |
recombinant His6-tagged enzyme, gel filtration | Vulcanisaeta moutnovskia |
Organism | UniProt | Comment | Textmining |
---|---|---|---|
Vulcanisaeta moutnovskia | F0QW25 | isolated from a hot spring in Kamchatka, Russia | - |
Vulcanisaeta moutnovskia 768-28 | F0QW25 | isolated from a hot spring in Kamchatka, Russia | - |
Oxidation Stability | Organism |
---|---|
the enzyme activity in the presence of 15% v/v organic solvents showes significant tolerance of VMUT0738 to solvents | Vulcanisaeta moutnovskia |
Purification (Comment) | Organism |
---|---|
recombinant His6-tagged enzyme from Escherichia coli strain DLT1270 by nickel affinity chromatography and gel filtration | Vulcanisaeta moutnovskia |
Source Tissue | Comment | Organism | Textmining |
---|---|---|---|
additional information | Vulcanisaeta moutnovskia strain 768-28 is an obligately anaerobic acidophile with a pH range of growth from pH 3.5 to 6.5 and a temperature range of growth from 60 to 98°C | Vulcanisaeta moutnovskia | - |
Specific Activity Minimum [µmol/min/mg] | Specific Activity Maximum [µmol/min/mg] | Comment | Organism |
---|---|---|---|
0.0076 | - |
substrate 2-methylbenzylamine, purified recombinant His-tagged enzyme, pH 8.0, 65°C | Vulcanisaeta moutnovskia |
0.23 | - |
substrate pyruvate, with valine, purified recombinant His-tagged enzyme, pH 8.0, 65°C | Vulcanisaeta moutnovskia |
0.83 | - |
substrate 3-methyl-2-oxovalerate, purified recombinant His-tagged enzyme, pH 8.0, 65°C | Vulcanisaeta moutnovskia |
1.1 | - |
substrate DL-tryptophan, purified recombinant His-tagged enzyme, pH 8.0, 65°C | Vulcanisaeta moutnovskia |
1.6 | - |
substrate L-cysteine, purified recombinant His-tagged enzyme, pH 8.0, 65°C | Vulcanisaeta moutnovskia |
1.6 | - |
substrate L-phenylalanine, purified recombinant His-tagged enzyme, pH 8.0, 65°C | Vulcanisaeta moutnovskia |
1.65 | - |
substrate 4-methyl-2-oxovalerate, purified recombinant His-tagged enzyme, pH 8.0, 65°C | Vulcanisaeta moutnovskia |
1.65 | - |
substrate L-norleucine, purified recombinant His-tagged enzyme, pH 8.0, 65°C | Vulcanisaeta moutnovskia |
1.72 | - |
substrate L-leucine, purified recombinant His-tagged enzyme, pH 8.0, 65°C | Vulcanisaeta moutnovskia |
1.76 | - |
substrate L-histidine, purified recombinant His-tagged enzyme, pH 8.0, 65°C | Vulcanisaeta moutnovskia |
1.76 | - |
substrate L-isoleucine, purified recombinant His-tagged enzyme, pH 8.0, 65°C | Vulcanisaeta moutnovskia |
1.88 | - |
substrate indole-3-pyruvate, purified recombinant His-tagged enzyme, pH 8.0, 65°C | Vulcanisaeta moutnovskia |
2.64 | - |
substrate L-arginine, purified recombinant His-tagged enzyme, pH 8.0, 65°C | Vulcanisaeta moutnovskia |
2.96 | - |
substrate L-norvaline, purified recombinant His-tagged enzyme, pH 8.0, 65°C | Vulcanisaeta moutnovskia |
2.97 | - |
substrate 2-aminobutyrate, purified recombinant His-tagged enzyme, pH 8.0, 65°C | Vulcanisaeta moutnovskia |
3 | - |
substrate L-valine, purified recombinant His-tagged enzyme, pH 8.0, 65°C | Vulcanisaeta moutnovskia |
3.1 | - |
substrate 2-oxobutyrate, purified recombinant His-tagged enzyme, pH 8.0, 65°C | Vulcanisaeta moutnovskia |
3.56 | - |
substrate L-threonine, purified recombinant His-tagged enzyme, pH 8.0, 65°C | Vulcanisaeta moutnovskia |
3.9 | - |
substrate L-ornithine, purified recombinant His-tagged enzyme, pH 8.0, 65°C | Vulcanisaeta moutnovskia |
3.93 | - |
substrate DL-lysine, purified recombinant His-tagged enzyme, pH 8.0, 65°C | Vulcanisaeta moutnovskia |
4.37 | - |
substrate L-methionine, purified recombinant His-tagged enzyme, pH 8.0, 65°C | Vulcanisaeta moutnovskia |
Substrates | Comment Substrates | Organism | Products | Comment (Products) | Rev. | Reac. |
---|---|---|---|---|---|---|
2-methylbenzylamine + 4-methyl-2-oxovalerate | - |
Vulcanisaeta moutnovskia | acetophenone + L-leucine | - |
r | |
2-methylbenzylamine + 4-methyl-2-oxovalerate | - |
Vulcanisaeta moutnovskia 768-28 | acetophenone + L-leucine | - |
r | |
2-oxo-3-indolylpropanoate + L-leucine | - |
Vulcanisaeta moutnovskia | L-tryptophan + 4-methyl-2-oxovalerate | - |
r | |
2-oxo-3-indolylpropanoate + L-leucine | - |
Vulcanisaeta moutnovskia 768-28 | L-tryptophan + 4-methyl-2-oxovalerate | - |
r | |
2-oxobutyrate + L-leucine | - |
Vulcanisaeta moutnovskia | 2-aminobutyrate + 4-methyl-2-oxovalerate | - |
r | |
2-oxobutyrate + L-leucine | - |
Vulcanisaeta moutnovskia 768-28 | 2-aminobutyrate + 4-methyl-2-oxovalerate | - |
r | |
3-methyl-2-oxopentanoate + L-leucine | - |
Vulcanisaeta moutnovskia | L-isoleucine + 4-methyl-2-oxovalerate | - |
r | |
3-methyl-2-oxopentanoate + L-leucine | - |
Vulcanisaeta moutnovskia 768-28 | L-isoleucine + 4-methyl-2-oxovalerate | - |
r | |
4-methyl-2-oxopentanoate + L-leucine | - |
Vulcanisaeta moutnovskia | L-leucine + 4-methyl-2-oxovalerate | - |
r | |
DL-lysine + 4-methyl-2-oxovalerate | - |
Vulcanisaeta moutnovskia | 2-oxo-6-aminohexanoate + L-leucine | - |
r | |
DL-tryptophan + pyruvate | - |
Vulcanisaeta moutnovskia | 2-oxo-3-indolylpropanoate + L-alanine | - |
r | |
L-alanine + 4-methyl-2-oxovalerate | - |
Vulcanisaeta moutnovskia | pyruvate + L-leucine | - |
r | |
L-cysteine + 4-methyl-2-oxovalerate | - |
Vulcanisaeta moutnovskia | 2-oxo-3-thiobutyrate + L-leucine | - |
r | |
L-isoleucine + 4-methyl-2-oxovalerate | - |
Vulcanisaeta moutnovskia | 3-methyl-2-oxopentanoate + L-leucine | - |
r | |
L-leucine + 4-methyl-2-oxovalerate | - |
Vulcanisaeta moutnovskia | 4-methyl-2-oxopentanoate + L-leucine | - |
r | |
L-methionine + 4-methyl-2-oxovalerate | - |
Vulcanisaeta moutnovskia | 4-methylsulfanyl-2-oxobutanoate + L-leucine | - |
r | |
L-norleucine + 4-methyl-2-oxovalerate | - |
Vulcanisaeta moutnovskia | 2-oxohexanoate + L-leucine | - |
r | |
L-norvaline + 4-methyl-2-oxovalerate | - |
Vulcanisaeta moutnovskia | 2-oxovalerate + L-leucine | - |
r | |
L-ornithine + 4-methyl-2-oxovalerate | - |
Vulcanisaeta moutnovskia | ? + L-leucine | - |
r | |
L-phenylalanine + 4-methyl-2-oxovalerate | - |
Vulcanisaeta moutnovskia | phenylpyruvate + L-leucine | - |
r | |
L-threonine + 4-methyl-2-oxovalerate | - |
Vulcanisaeta moutnovskia | 2-oxo-3-hydroxybutyrate + L-leucine | - |
r | |
L-valine + 4-methyl-2-oxovalerate | - |
Vulcanisaeta moutnovskia | 3-methyl-2-oxobutanoate + L-leucine | - |
r | |
additional information | enzyme VMUT0738 displays broad and atypical substrate specificity. It shows absence of activity toward acidic substrates, high activity toward basic ones, and low but detectable activity toward the (R)-enantiomer of 2-methylbenzylamine. VMUT0738 shows specificity for typical BCAT substrates, L-branched-chain amino acids (BCAAs) and corresponding keto acids. The same or higher level of activity is observed in the overall reaction with aromatic amino acids, methionine, threonine and 2-aminobuturic acid. But activity with 2-oxoglutarate, which is a typical BCAT amino acceptor, is not detected at a concentration range from 0.5 to 20 mM. VMUT0738 is not active with both the corresponding amino acid Glu and another acidic amino acid (Asp). On the contrary, the specific activity of VMUT0738 with the basic amino acids ornithine, lysine and arginine is high. Enzyme VMUT0738 shows high stereospecificity for L-amino acids, no preference for BCAAs, exhibits unusual substrate specificity toward substrates with a basic side chain, and has no activity toward substrates with an acidic side chain | Vulcanisaeta moutnovskia | ? | - |
- |
|
additional information | enzyme VMUT0738 displays broad and atypical substrate specificity. It shows absence of activity toward acidic substrates, high activity toward basic ones, and low but detectable activity toward the (R)-enantiomer of 2-methylbenzylamine. VMUT0738 shows specificity for typical BCAT substrates, L-branched-chain amino acids (BCAAs) and corresponding keto acids. The same or higher level of activity is observed in the overall reaction with aromatic amino acids, methionine, threonine and 2-aminobuturic acid. But activity with 2-oxoglutarate, which is a typical BCAT amino acceptor, is not detected at a concentration range from 0.5 to 20 mM. VMUT0738 is not active with both the corresponding amino acid Glu and another acidic amino acid (Asp). On the contrary, the specific activity of VMUT0738 with the basic amino acids ornithine, lysine and arginine is high. Enzyme VMUT0738 shows high stereospecificity for L-amino acids, no preference for BCAAs, exhibits unusual substrate specificity toward substrates with a basic side chain, and has no activity toward substrates with an acidic side chain | Vulcanisaeta moutnovskia 768-28 | ? | - |
- |
|
pyruvate + L-alanine | - |
Vulcanisaeta moutnovskia | L-alanine + pyruvate | - |
r | |
pyruvate + L-leucine | - |
Vulcanisaeta moutnovskia | L-alanine + 4-methyl-2-oxovalerate | - |
r |
Subunits | Comment | Organism |
---|---|---|
homotetramer | 4 * 36800, a dimer of dimers, recombinant His6-tagged enzyme, SDS-PAGE | Vulcanisaeta moutnovskia |
More | the catalytically competent unit of BCATs is a dimer | Vulcanisaeta moutnovskia |
Synonyms | Comment | Organism |
---|---|---|
BcaT | - |
Vulcanisaeta moutnovskia |
branched-chain amino acid aminotransferase | - |
Vulcanisaeta moutnovskia |
VMUT0738 | - |
Vulcanisaeta moutnovskia |
VMUT_0738 | locus name | Vulcanisaeta moutnovskia |
Temperature Optimum [°C] | Temperature Optimum Maximum [°C] | Comment | Organism |
---|---|---|---|
65 | 70 | assay at | Vulcanisaeta moutnovskia |
Temperature Minimum [°C] | Temperature Maximum [°C] | Comment | Organism |
---|---|---|---|
60 | 90 | activity of VMUT0738 increases with a rise in the temperature from 60°C to 90°C | Vulcanisaeta moutnovskia |
Temperature Stability Minimum [°C] | Temperature Stability Maximum [°C] | Comment | Organism |
---|---|---|---|
70 | - |
purified enzyme VMUT0738, 24 h, 73% of the initial activity remains | Vulcanisaeta moutnovskia |
Turnover Number Minimum [1/s] | Turnover Number Maximum [1/s] | Substrate | Comment | Organism | Structure |
---|---|---|---|---|---|
2.07 | - |
L-norvaline | pH 8.0, 65°C | Vulcanisaeta moutnovskia | |
2.1 | - |
L-valine | pH 8.0, 65°C | Vulcanisaeta moutnovskia | |
2.44 | - |
L-ornithine | pH 8.0, 65°C | Vulcanisaeta moutnovskia |
pH Optimum Minimum | pH Optimum Maximum | Comment | Organism |
---|---|---|---|
8 | - |
- |
Vulcanisaeta moutnovskia |
Cofactor | Comment | Organism | Structure |
---|---|---|---|
pyridoxal 5'-phosphate | PLP, dependent on | Vulcanisaeta moutnovskia |
General Information | Comment | Organism |
---|---|---|
evolution | branched-chain amino acid aminotransferases (BCATs) belong to fold-type IV class of PLP enzymes and are referred to as alpha-aminotransferases. Sequence alignment reveals two motifs (V/I)xLDxR and PFG(K/H)YL characteristic of BCATs from species of the related genera Vulcanisaeta, Pyrobaculum and Thermoproteus that might be responsible for the unique substrate recognition profile of the enzyme | Vulcanisaeta moutnovskia |
additional information | enzyme structure comparisons, overview | Vulcanisaeta moutnovskia |
kcat/KM Value [1/mMs-1] | kcat/KM Value Maximum [1/mMs-1] | Substrate | Comment | Organism | Structure |
---|---|---|---|---|---|
0.31 | - |
L-ornithine | pH 8.0, 65°C | Vulcanisaeta moutnovskia | |
1.8 | - |
L-valine | pH 8.0, 65°C | Vulcanisaeta moutnovskia | |
2.46 | - |
L-norvaline | pH 8.0, 65°C | Vulcanisaeta moutnovskia |