Cloned (Comment) | Organism |
---|---|
recombinant expression of His-tagged wild-type and mutant enzymes in Escherichia coli | Arabidopsis thaliana |
recombinant expression of His-tagged wild-type and mutant enzymes in Escherichia coli | Medicago truncatula |
Crystallization (Comment) | Organism |
---|---|
purified recombinant enzyme, hanging drop vapor diffusion method, mixing of 7.5-10 mg/ml protein with precipitant solution containing 100 mM succinic acid, pH 7.0, and 15% PEG3350, or 100 mM bicine, pH 9.0, 7% PEG 6000, and 3 mM MgCl2 for the Mg2+-bound enzyme, 2 weeks, 19°C, 20% glycerol or PEG400 as cryoprotectants, X-ray diffraction structure determination and analysis at 1.93 A and 1.83 A resolution, respectively, structure modelling. Growth of AtPPA1 crystal is strongly correlated with the progression of proteolysis | Arabidopsis thaliana |
purified recombinant enzyme, hanging drop vapor diffusion method, mixing of 7.7 mg/ml bromelain-treated protein with precipitant solution containing 1.6% Tacsimate, pH 5.0, 80 mM tribasic sodium citrate, pH 5.6, 12.8% PEG 3350, and 20% glycerol, 2 days, 19°C, X-ray diffraction structure determination and analysis at 1.84-2.89A resolution, structure modelling | Medicago truncatula |
Protein Variants | Comment | Organism |
---|---|---|
D103N | site-directed mutagenesis | Arabidopsis thaliana |
D103N | site-directed mutagenesis | Medicago truncatula |
D135N | site-directed mutagenesis | Arabidopsis thaliana |
D135N | site-directed mutagenesis | Medicago truncatula |
D98N | site-directed mutagenesis | Arabidopsis thaliana |
D98N | site-directed mutagenesis | Medicago truncatula |
additional information | construction of an N-truncated variant of AtPPA1 with residues 1-29 deleted (DELTA(1-29)) produced using construct pMCSG48-AtPPA1-DELTA(1-29) | Arabidopsis thaliana |
additional information | construction of the truncated version of MtPPA1-DELTA(1-30), which is not possible to express | Medicago truncatula |
KM Value [mM] | KM Value Maximum [mM] | Substrate | Comment | Organism | Structure |
---|---|---|---|---|---|
0.0106 | - |
diphosphate | pH 7.5, 22°C, recombinant wild-type enzyme | Medicago truncatula | |
0.0358 | - |
diphosphate | pH 7.5, 22°C, recombinant wild-type enzyme | Arabidopsis thaliana | |
0.0432 | - |
diphosphate | pH 7.5, 22°C, recombinant truncated mutant AtPPA1-DELTA(1-29) | Arabidopsis thaliana |
Localization | Comment | Organism | GeneOntology No. | Textmining |
---|---|---|---|---|
mitochondrion | the N-terminal peptide of immature AtPPA1 is mostly disordered. It can be cleaved during maturation. The cleaved peptide is a mitochondrial targeting signal | Arabidopsis thaliana | 5739 | - |
mitochondrion | the N-terminal peptide of immature MtPPA1 can be cleaved during maturation. The cleaved peptide is a mitochondrial targeting signal | Medicago truncatula | 5739 | - |
soluble | - |
Arabidopsis thaliana | - |
- |
soluble | - |
Medicago truncatula | - |
- |
Metals/Ions | Comment | Organism | Structure |
---|---|---|---|
Mg2+ | required, metal coordination and binding site | Medicago truncatula | |
Mg2+ | required, natural cofactor of AtPPA1, metal coordination and binding site | Arabidopsis thaliana |
Molecular Weight [Da] | Molecular Weight Maximum [Da] | Comment | Organism |
---|---|---|---|
132000 | - |
gel filtration | Arabidopsis thaliana |
138000 | - |
gel filtration | Medicago truncatula |
Natural Substrates | Organism | Comment (Nat. Sub.) | Natural Products | Comment (Nat. Pro.) | Rev. | Reac. |
---|---|---|---|---|---|---|
diphosphate + H2O | Arabidopsis thaliana | - |
2 phosphate | - |
? | |
diphosphate + H2O | Medicago truncatula | - |
2 phosphate | - |
? |
Organism | UniProt | Comment | Textmining |
---|---|---|---|
Arabidopsis thaliana | Q93V56 | - |
- |
Medicago truncatula | I3STR7 | - |
- |
Posttranslational Modification | Comment | Organism |
---|---|---|
proteolytic modification | the N-terminal peptide is autocatalytically cleaved | Medicago truncatula |
proteolytic modification | the N-terminal peptide is autocatalytically cleaved, presence of three N-terminal variants, truncated at Leu23, Ser25 and Leu26. The growth of AtPPA1 crystal iis strongly correlated with the progression of proteolysis | Arabidopsis thaliana |
Purification (Comment) | Organism |
---|---|
recombinant His-tagged wild-type and mutant enzymes from Escherichia coli by nickel affinity chromatography, dialysis, tag cleavage by TEV protease, followed by gel filtration and ultrafiltration | Arabidopsis thaliana |
recombinant His-tagged wild-type and mutant enzymes from Escherichia coli by nickel affinity chromatography, dialysis, tag cleavage by TEV protease, followed by gel filtration and ultrafiltration | Medicago truncatula |
Substrates | Comment Substrates | Organism | Products | Comment (Products) | Rev. | Reac. |
---|---|---|---|---|---|---|
diphosphate + H2O | - |
Arabidopsis thaliana | 2 phosphate | - |
? | |
diphosphate + H2O | - |
Medicago truncatula | 2 phosphate | - |
? |
Subunits | Comment | Organism |
---|---|---|
homohexamer | 6 * 24500, dimer of trimers, recombinant detagged enzyme, SDS-PAGE | Medicago truncatula |
homohexamer | 6 * 25000, crecombinant detagged enzyme, SDS-PAGE | Arabidopsis thaliana |
More | topology and conformation of the PPA1 subunits, comparison to the enzyme from Arabidospis thaliana | Medicago truncatula |
More | topology and conformation of the PPA1 subunits, comparison to the enzyme from Medicago truncatula. The N-terminal peptide of immature AtPPA1 is mostly disordered | Arabidopsis thaliana |
Synonyms | Comment | Organism |
---|---|---|
AtPPA1 | - |
Arabidopsis thaliana |
inorganic pyrophosphatase | - |
Arabidopsis thaliana |
inorganic pyrophosphatase | - |
Medicago truncatula |
MtPPA1 | - |
Medicago truncatula |
PPase | - |
Arabidopsis thaliana |
PPase | - |
Medicago truncatula |
Temperature Optimum [°C] | Temperature Optimum Maximum [°C] | Comment | Organism |
---|---|---|---|
22 | - |
assay at room temperature | Arabidopsis thaliana |
22 | - |
assay at room temperature | Medicago truncatula |
Turnover Number Minimum [1/s] | Turnover Number Maximum [1/s] | Substrate | Comment | Organism | Structure |
---|---|---|---|---|---|
5.3 | - |
diphosphate | pH 7.5, 22°C, recombinant wild-type enzyme | Arabidopsis thaliana | |
10.8 | - |
diphosphate | pH 7.5, 22°C, recombinant wild-type enzyme | Medicago truncatula | |
21.4 | - |
diphosphate | pH 7.5, 22°C, recombinant truncated mutant AtPPA1-DELTA(1-29) | Arabidopsis thaliana |
pH Optimum Minimum | pH Optimum Maximum | Comment | Organism |
---|---|---|---|
7.5 | - |
assay at | Arabidopsis thaliana |
7.5 | - |
assay at | Medicago truncatula |
General Information | Comment | Organism |
---|---|---|
additional information | AtPPA1 three-dimensional structure analysis and modelling, overview | Arabidopsis thaliana |
additional information | MtPPA1 three-dimensional structure analysis and modelling, overview | Medicago truncatula |
physiological function | inorganic diphosphatases (PPases), which hydrolyze inorganic diphosphate to phosphate in the presence of divalent metal cations, play a key role in maintaining phosphorus homeostasis in cells | Arabidopsis thaliana |
physiological function | inorganic diphosphatases (PPases), which hydrolyze inorganic diphosphate to phosphate in the presence of divalent metal cations, play a key role in maintaining phosphorus homeostasis in cells | Medicago truncatula |
kcat/KM Value [1/mMs-1] | kcat/KM Value Maximum [1/mMs-1] | Substrate | Comment | Organism | Structure |
---|---|---|---|---|---|
148.1 | - |
diphosphate | pH 7.5, 22°C, recombinant wild-type enzyme | Arabidopsis thaliana | |
495.4 | - |
diphosphate | pH 7.5, 22°C, recombinant truncated mutant AtPPA1-DELTA(1-29) | Arabidopsis thaliana | |
1019 | - |
diphosphate | pH 7.5, 22°C, recombinant wild-type enzyme | Medicago truncatula |