Cloned (Comment) | Organism |
---|---|
expression of wild-type and mutant enzymes in Escherichia coli TB1 as maltose binding fusion proteins | Sus scrofa |
Protein Variants | Comment | Organism |
---|---|---|
K321Q | site-directed mutagenesis, kinetics are similar to the wild-type enzyme | Sus scrofa |
R314Q | site-directed mutagenesis, 10fold increased Km for NADP+ compared to the wild-type enzyme | Sus scrofa |
R323Q | site-directed mutagenesis, kinetics are similar to the wild-type enzyme | Sus scrofa |
Y316F | site-directed mutagenesis, kinetics are similar to the wild-type enzyme | Sus scrofa |
Y316L | site-directed mutagenesis, 4fold increased Km for NADP+ compared to the wild-type enzyme | Sus scrofa |
Inhibitors | Comment | Organism | Structure |
---|---|---|---|
NAD+ | - |
Sus scrofa |
KM Value [mM] | KM Value Maximum [mM] | Substrate | Comment | Organism | Structure |
---|---|---|---|---|---|
0.0003 | - |
Mn2+ | pH 7.4, recombinant wild-type enzyme | Sus scrofa | |
0.0004 | - |
Mn2+ | pH 7.4, recombinant mutant Y316F and K323Q | Sus scrofa | |
0.0005 | - |
Mn2+ | pH 7.4, recombinant mutant K321Q | Sus scrofa | |
0.0059 | - |
isocitrate | pH 7.4, recombinant mutant Y316F | Sus scrofa | |
0.0062 | - |
isocitrate | pH 7.4, recombinant mutant K323Q | Sus scrofa | |
0.0074 | - |
isocitrate | pH 7.4, recombinant wild-type enzyme | Sus scrofa | |
0.0077 | - |
NADP+ | pH 7.4, recombinant mutant K323Q | Sus scrofa | |
0.008 | - |
NADP+ | pH 7.4, recombinant mutant Y316F and K321Q | Sus scrofa | |
0.009 | - |
NADP+ | pH 7.4, recombinant wild-type enzyme | Sus scrofa | |
0.01 | - |
isocitrate | pH 7.4, recombinant mutant K321Q | Sus scrofa | |
0.032 | - |
NADP+ | pH 7.4, recombinant mutant Y316L | Sus scrofa | |
0.092 | - |
NADP+ | pH 7.4, recombinant mutant R314Q | Sus scrofa |
Localization | Comment | Organism | GeneOntology No. | Textmining |
---|---|---|---|---|
mitochondrion | - |
Sus scrofa | 5739 | - |
Metals/Ions | Comment | Organism | Structure |
---|---|---|---|
Mn2+ | - |
Sus scrofa |
Molecular Weight [Da] | Molecular Weight Maximum [Da] | Comment | Organism |
---|---|---|---|
82000 | - |
recombinant mutant Y316F, dynamic light scattering | Sus scrofa |
86000 | - |
recombinant wild-type enzyme, dynamic light scattering | Sus scrofa |
87000 | - |
recombinant mutants K321Q and R314Q, dynamic light scattering | Sus scrofa |
97900 | - |
recombinant mutant Y316L, native PAGE at pH 5.5 | Sus scrofa |
99800 | - |
recombinant wild-type enzyme, native PAGE at pH 5.5 | Sus scrofa |
Organism | UniProt | Comment | Textmining |
---|---|---|---|
Sus scrofa | - |
- |
- |
Purification (Comment) | Organism |
---|---|
recombinant wild-type and mutant enzymes from Escherichia coli, to homogeneity by amylose affinity chromatography, thrombin digestion, and DEAE ion exchange chromatography | Sus scrofa |
Specific Activity Minimum [µmol/min/mg] | Specific Activity Maximum [µmol/min/mg] | Comment | Organism |
---|---|---|---|
7 | - |
purified recombinant mutant Y316L | Sus scrofa |
18.1 | - |
purified recombinant mutant R314Q | Sus scrofa |
38.6 | - |
purified recombinant wild-type enzyme | Sus scrofa |
38.7 | - |
purified recombinant mutant K323Q | Sus scrofa |
39.6 | - |
purified recombinant mutant K321Q | Sus scrofa |
40.3 | - |
purified recombinant mutant Y316F | Sus scrofa |
Substrates | Comment Substrates | Organism | Products | Comment (Products) | Rev. | Reac. |
---|---|---|---|---|---|---|
isocitrate + NADP+ | - |
Sus scrofa | 2-oxoglutarate + CO2 + NADPH + H+ | - |
? |
Subunits | Comment | Organism |
---|---|---|
dimer | residues Arg314, Tyr316, Lys321, and Arg323 are not involved in subunit interaction | Sus scrofa |
Synonyms | Comment | Organism |
---|---|---|
NADP-dependent isocitrate dehydrogenase | - |
Sus scrofa |
pH Optimum Minimum | pH Optimum Maximum | Comment | Organism |
---|---|---|---|
7.4 | - |
assay at | Sus scrofa |
Cofactor | Comment | Organism | Structure |
---|---|---|---|
NADP+ | Arg314 and the aromatic ring of Tyr316 are important for affinity to the cofactor and catalysis, interaction with 2'-phosphate group of NADP+ | Sus scrofa |
Ki Value [mM] | Ki Value maximum [mM] | Inhibitor | Comment | Organism | Structure |
---|---|---|---|---|---|
8 | - |
NAD+ | pH 7.4, versus NADPH, recombinant mutant R314Q | Sus scrofa | |
11 | - |
NAD+ | pH 7.4, versus NADPH, recombinant wild-type enzyme and mutant Y316L | Sus scrofa | |
12 | - |
NAD+ | pH 7.4, versus NADPH, recombinant mutant Y316F | Sus scrofa |