Activating Compound | Comment | Organism | Structure |
---|---|---|---|
acetyl-CoA | nonessential activator. Both acetyl-CoA and Mg2+ assist in coupling the MgATP-dependent carboxylation of biotin in the biotin carboxylase (BC) domain with pyruvate carboxylation in the carboxyl transferase (CT) domain. Absence of acetyl-CoA results in only 9% of fully activated enzyme. Acetyl-CoA also has a noticeable effect on the activity of the oxamate-induced decarboxylation of oxaloacetate but no effect on the rate of MgADP phosphorylation by carbamoyl phosphate | Rhizobium etli |
Cloned (Comment) | Organism |
---|---|
expressed in Escherichia coli as a His-tagged fusion protein | Rhizobium etli |
Crystallization (Comment) | Organism |
---|---|
crystal structures of mutant T882A pyruvate carboxylase are determined cocrystallized with phosphonoacetate and MgADP | Rhizobium etli |
Protein Variants | Comment | Organism |
---|---|---|
K1119Q | mutant that lacks tethered biotin. Addition of 10 mM biotin increases the kcat of MgATP hydrolysis to rates observed for wild-type RePC in the absence of free biotin. This rate increase, coupled with a 35fold decrease in the Km for MgATP, results in a nearly 1000fold increase in the catalytic efficiency of the mutant K1119Q RePC catalyzed reaction when 10 mM free biotin is added | Rhizobium etli |
T882A | T882 mutant is a tetrameric holoenzyme where positioning of the tethered biotin favors placement in the BC domain. Free biotin increases the kcat for both the wild-type and the T882A mutant RePC-catalyzed reactions without having a major effect on the Km for MgATP. Crystal structures of mutant T882A pyruvate carboxylase are determined cocrystallized with phosphonoacetate and MgADP | Rhizobium etli |
Inhibitors | Comment | Organism | Structure |
---|---|---|---|
Phosphonoacetate | - |
Rhizobium etli |
KM Value [mM] | KM Value Maximum [mM] | Substrate | Comment | Organism | Structure |
---|---|---|---|---|---|
additional information | - |
additional information | 0.5 mM Mg2+: Vmax (micromol/mg/min) 1.16, Ka (acetyl-CoA) 0.038 mM/1 mM Mg2+: Vmax (micromol/mg/min) 1.39, Ka (acetyl-CoA) 0.026 mM/1.5 mM Mg2+: Vmax (micromol/mg/min) 2.59, Ka (acetyl-CoA) 0.025 mM/3 mM Mg2+: Vmax (micromol/mg/min) 5.7, Ka (acetyl-CoA) 0.01 mM | Rhizobium etli | |
0.018 | - |
ATP | pH 7.5, 25°C, wild-type, 10 mM biotin, pyruvate carboxylation | Rhizobium etli | |
0.022 | - |
ATP | pH 7.5, 25°C, wild-type, no biotin, pyruvate carboxylation | Rhizobium etli | |
0.04 | - |
ATP | pH 7.5, 25°C, mutant K119Q, 10 mM biotin, pyruvate carboxylation | Rhizobium etli | |
0.054 | - |
ATP | pH 7.5, 25°C, mutant T882A, no biotin, pyruvate carboxylation | Rhizobium etli | |
0.09 | - |
ATP | pH 7.5, 25°C, mutant T882A, 10 mM biotin, pyruvate carboxylation | Rhizobium etli | |
0.1 | 2 | ADP | pH 7.5, 25°C, wild-type, reverse reaction | Rhizobium etli | |
0.145 | - |
ATP | pH 7.5, 25°C, wild-type, pyruvate carboxylation | Rhizobium etli | |
0.15 | - |
pyruvate | 5 mM Mg2+, pH 7.5, 25°C, wild-type, pyruvate carboxylation, Vmax: 3.98 micromol/mg/min | Rhizobium etli | |
0.15 | - |
pyruvate | pH 7.5, 25°C, wild-type, pyruvate carboxylation | Rhizobium etli | |
1.2 | - |
ATP | pH 7.5, 25°C, mutant K119Q, no biotin, pyruvate carboxylation | Rhizobium etli | |
2.65 | - |
pyruvate | 3 mM Mg2+, pH 7.5, 25°C, wild-type, pyruvate carboxylation, Vmax: 3.18 micromol/mg/min | Rhizobium etli | |
3 | - |
pyruvate | 1.5 mM Mg2+, pH 7.5, 25°C, wild-type, pyruvate carboxylation, Vmax: 1.41 micromol/mg/min | Rhizobium etli | |
3.1 | - |
pyruvate | 0.7 mM Mg2+, pH 7.5, 25°C, wild-type, pyruvate carboxylation, Vmax: 0.69 micromol/mg/min | Rhizobium etli | |
10.8 | - |
HCO3- | pH 7.5, 25°C, wild-type, pyruvate carboxylation | Rhizobium etli |
Metals/Ions | Comment | Organism | Structure |
---|---|---|---|
Mg2+ | essential activator. Both acetyl-CoA and Mg2+ assist in coupling the MgATP-dependent carboxylation of biotin in the biotin carboxylase (BC) domain with pyruvate carboxylation in the carboxyl transferase (CT) domain. High Mg2+ concentration (above 7 mM) inhibits pyruvate carboxylation and MgATP cleavage but no inhibition of MgADP phosphorylation reaction | Rhizobium etli |
Organism | UniProt | Comment | Textmining |
---|---|---|---|
Rhizobium etli | Q2K340 | - |
- |
Rhizobium etli CFN 42 | Q2K340 | - |
- |
Purification (Comment) | Organism |
---|---|
using Ni-NTA chromatography | Rhizobium etli |
Substrates | Comment Substrates | Organism | Products | Comment (Products) | Rev. | Reac. |
---|---|---|---|---|---|---|
ATP + pyruvate + HCO3- | - |
Rhizobium etli | ADP + phosphate + oxaloacetate | - |
r | |
ATP + pyruvate + HCO3- | - |
Rhizobium etli CFN 42 | ADP + phosphate + oxaloacetate | - |
r |
Synonyms | Comment | Organism |
---|---|---|
pyruvate carboxylase | - |
Rhizobium etli |
RePC | - |
Rhizobium etli |
Temperature Optimum [°C] | Temperature Optimum Maximum [°C] | Comment | Organism |
---|---|---|---|
25 | - |
assay at | Rhizobium etli |
Turnover Number Minimum [1/s] | Turnover Number Maximum [1/s] | Substrate | Comment | Organism | Structure |
---|---|---|---|---|---|
0.0013 | - |
ATP | pH 7.5, 25°C, mutant K119Q, no biotin, pyruvate carboxylation | Rhizobium etli | |
0.013 | - |
ADP | pH 7.5, 25°C, wild-type, reverse reaction | Rhizobium etli | |
0.04 | - |
ATP | pH 7.5, 25°C, mutant K119Q, 10 mM biotin, pyruvate carboxylation | Rhizobium etli | |
0.043 | - |
ATP | pH 7.5, 25°C, wild-type, no biotin, pyruvate carboxylation | Rhizobium etli | |
0.05 | - |
ATP | pH 7.5, 25°C, wild-type, 10 mM biotin, pyruvate carboxylation | Rhizobium etli | |
0.23 | - |
ATP | pH 7.5, 25°C, mutant T882A, no biotin, pyruvate carboxylation | Rhizobium etli | |
0.6 | - |
ATP | pH 7.5, 25°C, mutant T882A, 10 mM biotin, pyruvate carboxylation | Rhizobium etli | |
6.6 | - |
ATP | pH 7.5, 25°C, wild-type, pyruvate carboxylation | Rhizobium etli | |
7.33 | - |
pyruvate | pH 7.5, 25°C, wild-type, pyruvate carboxylation | Rhizobium etli | |
11.6 | - |
HCO3- | pH 7.5, 25°C, wild-type, pyruvate carboxylation | Rhizobium etli |
pH Optimum Minimum | pH Optimum Maximum | Comment | Organism |
---|---|---|---|
7.5 | - |
assay at | Rhizobium etli |
Cofactor | Comment | Organism | Structure |
---|---|---|---|
biotin | - |
Rhizobium etli |
kcat/KM Value [1/mMs-1] | kcat/KM Value Maximum [1/mMs-1] | Substrate | Comment | Organism | Structure |
---|---|---|---|---|---|
0.0011 | - |
ATP | pH 7.5, 25°C, mutant K119Q, no biotin, pyruvate carboxylation | Rhizobium etli | |
0.11 | - |
ADP | pH 7.5, 25°C, wild-type, reverse reaction | Rhizobium etli | |
1.08 | - |
HCO3- | pH 7.5, 25°C, wild-type, pyruvate carboxylation | Rhizobium etli | |
1.125 | - |
ATP | pH 7.5, 25°C, mutant K119Q, 10 mM biotin, pyruvate carboxylation | Rhizobium etli | |
1.91 | - |
ATP | pH 7.5, 25°C, wild-type, no biotin, pyruvate carboxylation | Rhizobium etli | |
2.78 | - |
ATP | pH 7.5, 25°C, wild-type, 10 mM biotin, pyruvate carboxylation | Rhizobium etli | |
4.33 | - |
ATP | pH 7.5, 25°C, mutant T882A, no biotin, pyruvate carboxylation | Rhizobium etli | |
7.16 | - |
ATP | pH 7.5, 25°C, mutant T882A, 10 mM biotin, pyruvate carboxylation | Rhizobium etli | |
44.5 | - |
ATP | pH 7.5, 25°C, wild-type, pyruvate carboxylation | Rhizobium etli | |
48.3 | - |
pyruvate | pH 7.5, 25°C, wild-type, pyruvate carboxylation | Rhizobium etli |