Cloned (Comment) | Organism |
---|---|
wild-type and mutants in pET-24d for expression in Escherichia coli BL21(DE3) | Escherichia coli |
Crystallization (Comment) | Organism |
---|---|
mutants P101S and P101L in complex with shikimate 3-phosphate and with or without inhibitor glyphosate, long-range structural change in glyphosate binding site compared to wild-type, Gly96 and Thr97 are shifted towards the glyphosate-binding site, which gets slightly narrowed, crystals: space group: P2(1)2(1)2(1), unit cell parameters: a: 57.6-57.8, b: 85.1-85.6, c: 87.9-88.3, beta: 90°, hanging-drop vapour-diffusion, protein solution (37.5 mg/ml, 5 mM inhibitor and/or 5 mM shikimate 3-phosphate), 19°C, molecular replacement using PDB: 1G6S as model | Escherichia coli |
Protein Variants | Comment | Organism |
---|---|---|
P101A | slight decrease in catalytic efficiency, decreased inhibitory potency of glyphosate | Escherichia coli |
P101G | slight decrease in catalytic efficiency, decreased inhibitory potency of glyphosate | Escherichia coli |
P101L | lowest catalytic efficiency, decreased inhibitory potency of glyphosate due to long-range conformational changes | Escherichia coli |
P101S | slight decrease in catalytic efficiency, decreased inhibitory potency of glyphosate due to long-range conformational changes, flexibility of Ser101 side chain disables hydrophobic bonding with carbonyl oxygen of Thr97 | Escherichia coli |
Inhibitors | Comment | Organism | Structure |
---|---|---|---|
glyphosate | - |
Escherichia coli |
KM Value [mM] | KM Value Maximum [mM] | Substrate | Comment | Organism | Structure |
---|---|---|---|---|---|
additional information | - |
phosphoenolpyruvate | KM/KI: 13, mutant P101S | Escherichia coli | |
additional information | - |
phosphoenolpyruvate | KM/KI: 150 | Escherichia coli | |
additional information | - |
phosphoenolpyruvate | KM/KI: 2.3, mutant P101L | Escherichia coli | |
additional information | - |
phosphoenolpyruvate | KM/KI: 4.2, mutant P101A | Escherichia coli | |
additional information | - |
phosphoenolpyruvate | KM/KI: 7.5, mutant P101G | Escherichia coli | |
additional information | - |
additional information | small residue substitutions for Pro101 do not alter the KM for phosphoenolpyruvate and shikimate 3-phosphate | Escherichia coli | |
0.06 | - |
shikimate 3-phosphate | +/-0.004 mM, mutant P101A | Escherichia coli | |
0.06 | - |
phosphoenolpyruvate | +/-0.006 mM | Escherichia coli | |
0.06 | - |
shikimate 3-phosphate | +/-0.006 mM | Escherichia coli | |
0.06 | - |
shikimate 3-phosphate | +/-0.010 mM, mutant P101G | Escherichia coli | |
0.07 | - |
phosphoenolpyruvate | +/-0.005 mM, mutant P101S | Escherichia coli | |
0.08 | - |
phosphoenolpyruvate | +/-0.005 mM, mutant P101A | Escherichia coli | |
0.08 | - |
shikimate 3-phosphate | +/-0.006 mM, mutant P101S | Escherichia coli | |
0.09 | - |
phosphoenolpyruvate | +/-0.010 mM, mutant P101G | Escherichia coli | |
0.1 | - |
shikimate 3-phosphate | +/-0.007 mM, mutant P101L | Escherichia coli | |
0.15 | - |
phosphoenolpyruvate | +/-0.020 mM, mutant P101L | Escherichia coli |
Organism | UniProt | Comment | Textmining |
---|---|---|---|
Escherichia coli | P0A6D3 | - |
- |
Specific Activity Minimum [µmol/min/mg] | Specific Activity Maximum [µmol/min/mg] | Comment | Organism |
---|---|---|---|
8 | - |
+/-0.2 micromol/min/mg, mutant P101L | Escherichia coli |
22 | - |
+/-0.4 micromol/min/mg, mutant P101S | Escherichia coli |
28 | - |
+/-1 micromol/min/mg, mutant P101G | Escherichia coli |
31 | - |
+/-0.5 micromol/min/mg, mutant P101A | Escherichia coli |
50 | - |
+/-1 micromol/min/mg | Escherichia coli |
Substrates | Comment Substrates | Organism | Products | Comment (Products) | Rev. | Reac. |
---|---|---|---|---|---|---|
shikimate 3-phosphate + phosphoenolpyruvate | 30 min, 25°C, pH 7.5, in presence of 2 mM dithiothreitol | Escherichia coli | phosphate + 5-enolpyruvylshikimate 3-phosphate | detection of phosphate generation with Lanzetta reagent and change in absorbance at 650 nm | ? |
Synonyms | Comment | Organism |
---|---|---|
5-enolpyruvylshikimate 3-phosphate synthase | - |
Escherichia coli |
E. coli EPSPS | - |
Escherichia coli |
EPSPS | - |
Escherichia coli |
Turnover Number Minimum [1/s] | Turnover Number Maximum [1/s] | Substrate | Comment | Organism | Structure |
---|---|---|---|---|---|
additional information | - |
shikimate 3-phosphate | kcat/KM: 200000 1/M*s, mutant P101S | Escherichia coli | |
additional information | - |
phosphoenolpyruvate | kcat/KM: 240000 1/M*s, mutant P101G | Escherichia coli | |
additional information | - |
phosphoenolpyruvate | kcat/KM: 280000 1/M*s, mutant P101A | Escherichia coli | |
additional information | - |
phosphoenolpyruvate | kcat/KM: 290000 1/M*s, mutant P101S | Escherichia coli | |
additional information | - |
shikimate 3-phosphate | kcat/KM: 390000 1/M*s, mutant P101A | Escherichia coli | |
additional information | - |
shikimate 3-phosphate | kcat/KM: 390000 1/M*s, mutant P101G | Escherichia coli | |
additional information | - |
phosphoenolpyruvate | kcat/KM: 41000 1/M*s, mutant P101L | Escherichia coli | |
additional information | - |
shikimate 3-phosphate | kcat/KM: 62000 1/M*s, mutant P101L | Escherichia coli | |
additional information | - |
phosphoenolpyruvate | kcat/KM: 640000 1/M*s | Escherichia coli | |
additional information | - |
shikimate 3-phosphate | kcat/KM: 640000 1/M*s | Escherichia coli |
Ki Value [mM] | Ki Value maximum [mM] | Inhibitor | Comment | Organism | Structure |
---|---|---|---|---|---|
additional information | - |
glyphosate | any substitutions for residue Pro101 decrease glyphosate binding affinity and inhibitor potency | Escherichia coli | |
0.0004 | - |
glyphosate | +/-0.00006 mM | Escherichia coli | |
0.0055 | - |
glyphosate | +/-0.0003 mM, mutant P101S | Escherichia coli | |
0.012 | - |
glyphosate | +/-0.004 mM, mutant P101G | Escherichia coli | |
0.019 | - |
glyphosate | +/-0.003 mM, mutant P101A | Escherichia coli | |
0.066 | - |
glyphosate | +/-0.002 mM, mutant P101L | Escherichia coli |