Application | Comment | Organism |
---|---|---|
additional information | phylogenetically variant active-site residues are useful for modulating activity of transketolase on natural or structurally-homologous substrates, whereas conserved residues which no longer interact with modified target substrates are useful sites to apply saturation mutagenesis for improvement of activity of transketolase | Escherichia coli |
Cloned (Comment) | Organism |
---|---|
tkta gene complete with its natural promoter on the pQR711 plasmid expressed in Escherichia coli TOP10 or XL10 cells. A29E mutation introduced into the pQR412 vector, in which the transketolase gene has an N-terminal His×6 tag. Both wild-type and mutant A29E overexpressed from pQR412 | Escherichia coli |
Protein Variants | Comment | Organism |
---|---|---|
A29D | specific activity is 2.7fold greater than for the wild-type | Escherichia coli |
A29E | specific activity is 3fold greater than for the wild-type | Escherichia coli |
D259A | specific activity is 2.3fold greater than for the wild-type | Escherichia coli |
D259G | specific activity is 2fold greater than for the wild-type | Escherichia coli |
H461S | specific activity is 4.8fold greater than for the wild-type | Escherichia coli |
R358I | specific activity is 2.1fold greater than for the wild-type | Escherichia coli |
R358P | specific activity is 1.5fold greater than for the wild-type | Escherichia coli |
R520G | specific activity is 2.1fold greater than for the wild-type | Escherichia coli |
R520P | specific activity is 2.3fold greater than for the wild-type | Escherichia coli |
R520Stop | specific activity is 3.6fold greater than for the wild-type | Escherichia coli |
R520V | specific activity is 3.6fold greater than for the wild-type | Escherichia coli |
S188Q | specific activity is 2.3fold greater than for the wild-type | Escherichia coli |
KM Value [mM] | KM Value Maximum [mM] | Substrate | Comment | Organism | Structure |
---|---|---|---|---|---|
5.3 | - |
Hydroxypyruvate | wild-type, in the presence of 50 mM glycolaldehyde | Escherichia coli | |
6.6 | - |
Hydroxypyruvate | mutant A29E, in the presence of 50 mM glycolaldehyde | Escherichia coli | |
35 | - |
glycolaldehyde | wild-type, in the presence of 50 mM hydroxypyruvate | Escherichia coli | |
200 | - |
glycolaldehyde | mutant A29E, in the presence of 50 mM hydroxypyruvate | Escherichia coli |
Organism | UniProt | Comment | Textmining |
---|---|---|---|
Escherichia coli | - |
- |
- |
Purification (Comment) | Organism |
---|---|
wild-type and mutant A29E purified on Ni-NTA resin | Escherichia coli |
Specific Activity Minimum [µmol/min/mg] | Specific Activity Maximum [µmol/min/mg] | Comment | Organism |
---|---|---|---|
0.65 | - |
wild-type, in the presence of 50 mM hydroxypyruvate, 50 mM glycolaldehyde, and 50 mM Tris-HCl, 2.4 mM thiamine diphosphate, 9 mM MgCl2, pH 7.0 | Escherichia coli |
1 | - |
mutant R358P, in the presence of 50 mM hydroxypyruvate, 50 mM glycolaldehyde, and 50 mM Tris-HCl, 2.4 mM thiamine diphosphate, 9 mM MgCl2, pH 7.0 | Escherichia coli |
1.3 | - |
mutant D259G, in the presence of 50 mM hydroxypyruvate, 50 mM glycolaldehyde, and 50 mM Tris-HCl, 2.4 mM thiamine diphosphate, 9 mM MgCl2, pH 7.0 | Escherichia coli |
1.37 | - |
mutant R358I, in the presence of 50 mM hydroxypyruvate, 50 mM glycolaldehyde, and 50 mM Tris-HCl, 2.4 mM thiamine diphosphate, 9 mM MgCl2, pH 7.0 | Escherichia coli |
1.4 | - |
mutant R520G, in the presence of 50 mM hydroxypyruvate, 50 mM glycolaldehyde, and 50 mM Tris-HCl, 2.4 mM thiamine diphosphate, 9 mM MgCl2, pH 7.0 | Escherichia coli |
1.5 | - |
mutant D259A, in the presence of 50 mM hydroxypyruvate, 50 mM glycolaldehyde, and 50 mM Tris-HCl, 2.4 mM thiamine diphosphate, 9 mM MgCl2, pH 7.0 | Escherichia coli |
1.5 | - |
mutant R520P, in the presence of 50 mM hydroxypyruvate, 50 mM glycolaldehyde, and 50 mM Tris-HCl, 2.4 mM thiamine diphosphate, 9 mM MgCl2, pH 7.0 | Escherichia coli |
1.5 | - |
mutant S188Q, in the presence of 50 mM hydroxypyruvate, 50 mM glycolaldehyde, and 50 mM Tris-HCl, 2.4 mM thiamine diphosphate, 9 mM MgCl2, pH 7.0 | Escherichia coli |
1.8 | - |
mutant A29D, in the presence of 50 mM hydroxypyruvate, 50 mM glycolaldehyde, and 50 mM Tris-HCl, 2.4 mM thiamine diphosphate, 9 mM MgCl2, pH 7.0 | Escherichia coli |
1.95 | - |
mutant A29E, in the presence of 50 mM hydroxypyruvate, 50 mM glycolaldehyde, and 50 mM Tris-HCl, 2.4 mM thiamine diphosphate, 9 mM MgCl2, pH 7.0 | Escherichia coli |
2.3 | - |
mutant R520Stop, in the presence of 50 mM hydroxypyruvate, 50 mM glycolaldehyde, and 50 mM Tris-HCl, 2.4 mM thiamine diphosphate, 9 mM MgCl2, pH 7.0 | Escherichia coli |
2.3 | - |
mutant R520V, in the presence of 50 mM hydroxypyruvate, 50 mM glycolaldehyde, and 50 mM Tris-HCl, 2.4 mM thiamine diphosphate, 9 mM MgCl2, pH 7.0 | Escherichia coli |
3.14 | - |
mutant H461S, in the presence of 50 mM hydroxypyruvate, 50 mM glycolaldehyde, and 50 mM Tris-HCl, 2.4 mM thiamine diphosphate, 9 mM MgCl2, pH 7.0 | Escherichia coli |
Substrates | Comment Substrates | Organism | Products | Comment (Products) | Rev. | Reac. |
---|---|---|---|---|---|---|
hydroxypyruvate + glycolaldehyde | - |
Escherichia coli | L-erythrulose + ? | - |
? |
Synonyms | Comment | Organism |
---|---|---|
TktA | - |
Escherichia coli |
Cofactor | Comment | Organism | Structure |
---|---|---|---|
thiamine diphosphate | - |
Escherichia coli |