Application | Comment | Organism |
---|---|---|
synthesis | enzyme cascade from threonine to synthesis of L-2-aminobutanoate. In this cascade, the threonine deaminase is used for threonine to 2-oxobutanoate, then LeuDH mutant and formate dehydrogenase are used for synthesis of L-2-aminobutanoate. Under optimized conditions, 1 M threonine is catalyzed by whole cells of Escherichia coli harboring the enzymes in 12 h in sodium phosphate buffer to the optically pure L-2-aminobutanoate with a yield of 99% and ee above 99% | Bacillus cereus |
Crystallization (Comment) | Organism |
---|---|
homology modeling of structure and semirational engineering to increase the catalytic efficiency | Bacillus cereus |
modeling of structure. Active site residues Lys81, Asp116, and Lys69 are catalytically important. Lys81 linkes the -OH group of the substrate 2-oxobutanoate by a 2.3-A long H-bond, and Asp116 is involved in proton transfer during catalysis | Bacillus cereus |
Protein Variants | Comment | Organism |
---|---|---|
M347G | 251% of wild-type activity | Bacillus cereus |
M347G | 341% of wild-type activity | Bacillus cereus |
M347G/Q358N | 400% of wild-type activity | Bacillus cereus |
M347N | 228% of wild-type activity | Bacillus cereus |
N262N | 121% of wild-type activity | Bacillus cereus |
N70F | 128% of wild-type activity | Bacillus cereus |
Q358N | 310% of wild-type activity | Bacillus cereus |
Q358N | 459% of wild-type activity | Bacillus cereus |
Q358T | 248% of wild-type activity | Bacillus cereus |
Q358T | 257% of wild-type activity | Bacillus cereus |
KM Value [mM] | KM Value Maximum [mM] | Substrate | Comment | Organism | Structure |
---|---|---|---|---|---|
13.5 | - |
2-oxobutanoate | mutant Q358N, pH 7.5, 37°C | Bacillus cereus | |
13.53 | - |
2-oxobutanoate | mutant Q358N, pH 7.5, 37°C | Bacillus cereus | |
16.8 | - |
2-oxobutanoate | mutant M347G/Q358N, pH 7.5, 37°C | Bacillus cereus | |
21.3 | - |
2-oxobutanoate | mutant M374G, pH 7.5, 37°C | Bacillus cereus | |
21.33 | - |
2-oxobutanoate | mutant M347G, pH 7.5, 37°C | Bacillus cereus | |
22.7 | - |
2-oxobutanoate | wild-type, pH 7.5, 37°C | Bacillus cereus | |
22.74 | - |
2-oxobutanoate | wild-type, pH 7.5, 37°C | Bacillus cereus |
Organism | UniProt | Comment | Textmining |
---|---|---|---|
Bacillus cereus | - |
- |
- |
Specific Activity Minimum [µmol/min/mg] | Specific Activity Maximum [µmol/min/mg] | Comment | Organism |
---|---|---|---|
2.6 | - |
wild-type, pH 7.5, 37°C | Bacillus cereus |
8.88 | - |
mutant M374G, pH 7.5, 37°C | Bacillus cereus |
10.4 | - |
mutant M347G/Q358N, pH 7.5, 37°C | Bacillus cereus |
11.94 | - |
mutant Q358N, pH 7.5, 37°C | Bacillus cereus |
Substrates | Comment Substrates | Organism | Products | Comment (Products) | Rev. | Reac. |
---|---|---|---|---|---|---|
2-oxobutanoate + NH3 + NADH + H+ | - |
Bacillus cereus | L-2-aminobutanoate + H2O + NAD+ | - |
? | |
2-oxobutanoate + NH3 + NADH + H+ | - |
Bacillus cereus | L-2-aminobutanoate + H2O + NAD+ | - |
r | |
L-leucine + H2O + NAD+ | - |
Bacillus cereus | 4-methyl-2-oxopentanoate + NH3 + NADH + H+ | - |
r |
Turnover Number Minimum [1/s] | Turnover Number Maximum [1/s] | Substrate | Comment | Organism | Structure |
---|---|---|---|---|---|
0.055 | - |
2-oxobutanoate | wild-type, pH 7.5, 37°C | Bacillus cereus | |
0.056 | - |
2-oxobutanoate | wild-type, pH 7.5, 37°C | Bacillus cereus | |
0.15 | - |
2-oxobutanoate | mutant Q358N, pH 7.5, 37°C | Bacillus cereus | |
0.46 | - |
2-oxobutanoate | mutant M347G, pH 7.5, 37°C | Bacillus cereus | |
0.461 | - |
2-oxobutanoate | mutant M374G, pH 7.5, 37°C | Bacillus cereus | |
0.826 | - |
2-oxobutanoate | mutant M347G/Q358N, pH 7.5, 37°C | Bacillus cereus | |
0.83 | - |
2-oxobutanoate | mutant M347G/Q358N, pH 7.5, 37°C | Bacillus cereus | |
1.5 | - |
2-oxobutanoate | mutant Q358N, pH 7.5, 37°C | Bacillus cereus |
kcat/KM Value [1/mMs-1] | kcat/KM Value Maximum [1/mMs-1] | Substrate | Comment | Organism | Structure |
---|---|---|---|---|---|
0.0025 | - |
2-oxobutanoate | wild-type, pH 7.5, 37°C | Bacillus cereus | |
0.0217 | - |
2-oxobutanoate | mutant M374G, pH 7.5, 37°C | Bacillus cereus | |
0.022 | - |
2-oxobutanoate | mutant M347G, pH 7.5, 37°C | Bacillus cereus | |
0.0492 | - |
2-oxobutanoate | mutant M347G/Q358N, pH 7.5, 37°C | Bacillus cereus | |
0.05 | - |
2-oxobutanoate | mutant M347G/Q358N, pH 7.5, 37°C | Bacillus cereus | |
0.11 | - |
2-oxobutanoate | mutant Q358N, pH 7.5, 37°C | Bacillus cereus | |
0.1112 | - |
2-oxobutanoate | mutant Q358N, pH 7.5, 37°C | Bacillus cereus |