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Literature summary for 2.7.1.40 extracted from
- The contribution of two isozymes to the pyruvate kinase activity of Vibrio cholerae one K+-dependent constitutively active and another K+-independent with essential allosteric activation (2017), PLoS ONE, 12, e0178673 .
Activating Compound
| Activating Compound | Comment | Organism | Structure |
|---|---|---|---|
| AMP | slight activation | Vibrio cholerae serotype O1 |  |
| fructose 1,6-bisphosphate | slight activation | Vibrio cholerae serotype O1 | |
| fructose 1,6-bisphosphate | slight activation, the isozyme with Glu117 is an active K+-dependent enzyme, at the same substrate concentration, its Vmax in the absence of fructose 1,6-bisphosphate is 80% of that with its effector. VcIPK is activated 31fold by fructose 1,6-bisphosphate | Vibrio cholerae serotype O1 | |
| fructose 2,6-bisphosphate | - |
Vibrio cholerae serotype O1 | |
| fructose 6-phosphate | slight activation | Vibrio cholerae serotype O1 | |
| fructose 6-phosphate | moderate activation | Vibrio cholerae serotype O1 | |
| glucose 6-phosphate | slight activation | Vibrio cholerae serotype O1 | |
| glucose 6-phosphate | VcIIPK is activated 159fold by glucose 6-phosphate | Vibrio cholerae serotype O1 | |
| ribose 5-phosphate | slight activation | Vibrio cholerae serotype O1 | |
| ribose 5-phosphate | Rib 5-P, VcIIPK is activated 200fold by ribose 5-phosphate. the pyruvate kinase with Lys117 is a K+-independent enzyme displaying an allosteric activation by ribose 5-phosphate. In the K+-independent enzyme, Mn2+ may mimic the allosteric effect of ribose 5-phosphate | Vibrio cholerae serotype O1 | |
| ribose 5-phosphate | the pyruvate kinase with Lys117 is a K+-independent enzyme displaying an allosteric activation by ribose 5-phosphate. In the K+-independent enzyme, Mn2+ may mimic the allosteric effect of ribose 5-phosphate | Vibrio cholerae serotype O1 | |
Cloned(Commentary)
| Cloned (Comment) | Organism |
|---|---|
| recombinant expression of His-tagged isozyme PKI from pMCSG7/VcIPK plasmid in Escherichia coli strain BL21-CodonPlus (DE3)-RIL | Vibrio cholerae serotype O1 |
| recombinant expression of His-tagged isozyme PKII from pMCSG7/VcIIPK plasmid in Escherichia coli strain BL21-CodonPlus (DE3)-RIL | Vibrio cholerae serotype O1 |
Inhibitors
| Inhibitors | Comment | Organism | Structure |
|---|---|---|---|
| ADP-Cr2+ | a dead-end inhibitor; a dead-end inhibitor | Vibrio cholerae serotype O1 | |
| oxalate | a dead-end inhibitor; a dead-end inhibitor | Vibrio cholerae serotype O1 | |
KM Value [mM]
| KM Value [mM] | KM Value Maximum [mM] | Substrate | Comment | Organism | Structure |
|---|---|---|---|---|---|
| additional information | - |
additional information | despite their different allosteric behavior, both isozymes display a rapid equilibrium random order kinetic mechanism. Kinetic analysis, detailed overview | Vibrio cholerae serotype O1 | |
| 0.06 | - |
phosphoenolpyruvate | pH 7.0, 25°C, recombinant non-His-tagged enzyme, in presence of fructose 1,6-bisphosphate | Vibrio cholerae serotype O1 | |
| 0.069 | - |
phosphoenolpyruvate | pH 7.0, 25°C, recombinant His-tagged enzyme, in presence of fructose 1,6-bisphosphate | Vibrio cholerae serotype O1 | |
| 0.26 | - |
ADP | pH 7.0, 25°C, recombinant His-tagged enzyme | Vibrio cholerae serotype O1 | |
| 0.27 | - |
ADP | pH 7.0, 25°C, recombinant non-His-tagged enzyme | Vibrio cholerae serotype O1 | |
| 0.55 | - |
phosphoenolpyruvate | pH 7.0, 25°C, recombinant His-tagged enzyme | Vibrio cholerae serotype O1 | |
| 0.68 | - |
phosphoenolpyruvate | pH 7.0, 25°C, recombinant non-His-tagged enzyme | Vibrio cholerae serotype O1 | |
| 1 | - |
phosphoenolpyruvate | pH 7.0, 25°C, recombinant His-tagged enzyme, in presence of glucose 6-phosphate | Vibrio cholerae serotype O1 | |
Metals/Ions
| Metals/Ions | Comment | Organism | Structure |
|---|---|---|---|
| Cs+ | activates, at saturating concentrations of PEP3-, ADP-Mg complex and free Mg2+, the Km for Cs+ is 13.5 mM | Vibrio cholerae serotype O1 | |
| K+ | K117 isozyme requires K+ | Vibrio cholerae serotype O1 | |
| K+ | K117 isozyme requires K+, at saturating concentrations of PEP3-, ADP-Mg complex and free Mg2+, the Km for K+ is 8.5 mM | Vibrio cholerae serotype O1 | |
| Mg2+ | required for activity | Vibrio cholerae serotype O1 | |
| Mn2+ | the K+-independent enzyme, Mn2+ may mimic the allosteric effect of ribose 5-phosphate | Vibrio cholerae serotype O1 | |
| additional information | the pyruvate kinase isozyme sequences with Glu117 have been found to be K+-dependent, whereas those with Lys117 are K+-independent | Vibrio cholerae serotype O1 | |
| additional information | the pyruvate kinase isozyme sequences with Glu117 have been found to be K+-dependent, whereas those with Lys117 are K+-independent. In comparison with other K+-dependent PKs, VcIPK exhibits higher kcat and 2-5fold lower Km for the monovalent cations. No activation by Na+ and Li+. VCIPK is a good example of type Ib activation by monovalent cations. In such a mechanism, M+ coordination is absolutely required for catalysis, where both kcat and kcat/Km increase hyperbolically with [K+] | Vibrio cholerae serotype O1 | |
| NH4+ | activates, at saturating concentrations of PEP3-, ADP-Mg complex and free Mg2+, the Km for NH4+ is 2.7 mM | Vibrio cholerae serotype O1 | |
| Rb+ | activates, at saturating concentrations of PEP3-, ADP-Mg complex and free Mg2+, the Km for Rb+ is 5.9 mM | Vibrio cholerae serotype O1 | |
Molecular Weight [Da]
| Molecular Weight [Da] | Molecular Weight Maximum [Da] | Comment | Organism |
|---|---|---|---|
| 200000 | - |
recombinant His-tagged VcIIPK, native PAGE | Vibrio cholerae serotype O1 |
| 200000 | - |
recombinant His-tagged VcIPK, native PAGE | Vibrio cholerae serotype O1 |
Natural Substrates/ Products (Substrates)
| Natural Substrates | Organism | Comment (Nat. Sub.) | Natural Products | Comment (Nat. Pro.) | Rev. | Reac. |
|---|---|---|---|---|---|---|
| ADP + phosphoenolpyruvate | Vibrio cholerae serotype O1 | - |
ATP + pyruvate | - |
? | |
| ADP + phosphoenolpyruvate | Vibrio cholerae serotype O1 El Tor Inaba N16961 | - |
ATP + pyruvate | - |
? | |
| ADP + phosphoenolpyruvate | Vibrio cholerae serotype O1 ATCC 39315 | - |
ATP + pyruvate | - |
? | |
Organism
| Organism | UniProt | Comment | Textmining |
|---|---|---|---|
| Vibrio cholerae serotype O1 | Q9KQJ0 | - |
- |
| Vibrio cholerae serotype O1 | Q9KUN0 | - |
- |
| Vibrio cholerae serotype O1 ATCC 39315 | Q9KQJ0 | - |
- |
| Vibrio cholerae serotype O1 ATCC 39315 | Q9KUN0 | - |
- |
| Vibrio cholerae serotype O1 El Tor Inaba N16961 | Q9KQJ0 | - |
- |
| Vibrio cholerae serotype O1 El Tor Inaba N16961 | Q9KUN0 | - |
- |
Purification (Commentary)
| Purification (Comment) | Organism |
|---|---|
| recombinant His-tagged isozyme PKI from Escherichia coli strain BL21-CodonPlus (DE3)-RIL by nickel affinity chromatography and ammonium sulfate fraction, the tag cannot be cleaved, because the cleavage site is not accessible to the TEV protease, to 90% purity | Vibrio cholerae serotype O1 |
| recombinant His-tagged isozyme PKII from Escherichia coli strain BL21-CodonPlus (DE3)-RIL by nickel affinity chromatography and ammonium sulfate fraction, the tag cannot be cleaved, because the cleavage site is not accessible to the TEV protease, to 88% purity | Vibrio cholerae serotype O1 |
Source Tissue
| Source Tissue | Comment | Organism | Textmining |
|---|
Substrates and Products (Substrate)
| Substrates | Comment Substrates | Organism | Products | Comment (Products) | Rev. | Reac. |
|---|---|---|---|---|---|---|
| ADP + phosphoenolpyruvate | - |
Vibrio cholerae serotype O1 | ATP + pyruvate | - |
? | |
| ADP + phosphoenolpyruvate | - |
Vibrio cholerae serotype O1 El Tor Inaba N16961 | ATP + pyruvate | - |
? | |
| ADP + phosphoenolpyruvate | - |
Vibrio cholerae serotype O1 ATCC 39315 | ATP + pyruvate | - |
? | |
Subunits
| Subunits | Comment | Organism |
|---|---|---|
| homotetramer | 4 * 50000, recombinant His-tagged VcIIPK, SDS-PAGE, 4 * 54914, recombinant His-tagged VcIIPK, mass spectrometry, 4 * 52333, VcIIPK, sequence calculation | Vibrio cholerae serotype O1 |
| homotetramer | 4 * 50000, recombinant His-tagged VcIPK, SDS-PAGE, 4 * 53138, recombinant His-tagged VcIPK, mass spectrometry, 4 * 50439, VcIPK, sequence calculation | Vibrio cholerae serotype O1 |
Synonyms
| Synonyms | Comment | Organism |
|---|---|---|
| K+-dependent PK | - |
Vibrio cholerae serotype O1 |
| K+-independent PK | - |
Vibrio cholerae serotype O1 |
| VcIIPK | - |
Vibrio cholerae serotype O1 |
| VcIPK | - |
Vibrio cholerae serotype O1 |
| VC_0485 | - |
Vibrio cholerae serotype O1 |
| VC_2008 | - |
Vibrio cholerae serotype O1 |
Temperature Optimum [°C]
| Temperature Optimum [°C] | Temperature Optimum Maximum [°C] | Comment | Organism |
|---|---|---|---|
| 25 | - |
assay at | Vibrio cholerae serotype O1 |
pH Optimum
| pH Optimum Minimum | pH Optimum Maximum | Comment | Organism |
|---|---|---|---|
| 7 | - |
assay at | Vibrio cholerae serotype O1 |
Ki Value [mM]
| Ki Value [mM] | Ki Value maximum [mM] | Inhibitor | Comment | Organism | Structure |
|---|---|---|---|---|---|
| 0.0078 | - |
oxalate | pH 7.0, 25°C, recombinant non-His-tagged enzyme | Vibrio cholerae serotype O1 | |
| 0.071 | - |
oxalate | pH 7.0, 25°C, recombinant non-His-tagged enzyme | Vibrio cholerae serotype O1 | |
| 1.1 | - |
ADP-Cr2+ | pH 7.0, 25°C, recombinant non-His-tagged enzyme | Vibrio cholerae serotype O1 | |
| 1.3 | - |
ADP-Cr2+ | pH 7.0, 25°C, recombinant non-His-tagged enzyme | Vibrio cholerae serotype O1 | |
General Information
| General Information | Comment | Organism |
|---|---|---|
| physiological function | Vibrio cholerae has two isozymes that contribute to the pyruvate kinase activity: one K+-dependent constitutively active isozyme and another K+-independent isozyme with essential allosteric activation. The pyruvate kinase isozyme sequences with Glu117 have been found to be K+-dependent, whereas those with Lys117 are K+-independent | Vibrio cholerae serotype O1 |
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