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Literature summary for 1.1.1.42 extracted from

  • Kim, T.K.; Lee, P.; Colman, R.F.
    Critical role of Lys212 and Tyr140 in porcine NADP-dependent isocitrate dehydrogenase (2003), J. Biol. Chem., 278, 49323-49331.
    View publication on PubMed

Activating Compound

Activating Compound Comment Organism Structure
acetic acid mutant Y140T, 106fold activation compared to the wild-type enzyme Sus scrofa
ethylamine mutant K212Q, 4fold activation compared to the wild-type enzyme Sus scrofa
phenol mutant Y140T, 88fold activation compared to the wild-type enzyme Sus scrofa

Cloned(Commentary)

Cloned (Comment) Organism
expression of wild-type and mutant enzymes in Escherichia coli Sus scrofa

Protein Variants

Protein Variants Comment Organism
K212Q site-directed mutagenesis, highly decreased activity in both reaction directions compared to the wild-type enzyme, altered pH-dependency of the activity Sus scrofa
K212R site-directed mutagenesis, highly decreased activity in both reaction directions compared to the wild-type enzyme, altered pH-dependency of the activity Sus scrofa
K212Y site-directed mutagenesis, highly decreased activity in both reaction directions compared to the wild-type enzyme, altered pH-dependency of the activity Sus scrofa
Y140E site-directed mutagenesis, highly decreased activity in both reaction directions compared to the wild-type enzyme, unaltered Km for isocitrate and NADP+ Sus scrofa
Y140F site-directed mutagenesis, highly decreased activity in both reaction directions compared to the wild-type enzyme, unaltered Km for isocitrate and NADP+ Sus scrofa
Y140K site-directed mutagenesis, highly decreased activity in both reaction directions compared to the wild-type enzyme, unaltered Km for isocitrate and NADP+ Sus scrofa
Y140T site-directed mutagenesis, highly decreased activity in both reaction directions compared to the wild-type enzyme, unaltered Km for isocitrate and NADP+, highly increased activation by added exogenous acetic acid and phenol compared to the wild-type enzyme Sus scrofa

KM Value [mM]

KM Value [mM] KM Value Maximum [mM] Substrate Comment Organism Structure
0.00004
-
Mn2+ pH 7.4, mutant Y140F Sus scrofa
0.000084
-
Mn2+ pH 7.4, mutant Y140E Sus scrofa
0.00011
-
Mn2+ pH 7.4, wild-type enzyme Sus scrofa
0.00042
-
NADP+ pH 7.4, mutant Y140E Sus scrofa
0.00046
-
NADP+ pH 7.4, mutant Y140T Sus scrofa
0.00051
-
NADP+ pH 7.4, mutant Y140K Sus scrofa
0.0006
-
isocitrate pH 7.4, mutant Y140T Sus scrofa
0.0007
-
isocitrate pH 7.4, mutant Y140E Sus scrofa
0.00091
-
Mn2+ pH 7.4, mutant K212Q Sus scrofa
0.0011
-
Mn2+ pH 7.4, mutant K212Y Sus scrofa
0.0023
-
NADP+ pH 7.4, mutant Y140F Sus scrofa
0.0052
-
isocitrate pH 7.4, mutant Y140F Sus scrofa
0.0053
-
isocitrate pH 7.4, mutant Y140K Sus scrofa
0.0073
-
NADP+ pH 7.4, mutant K212Y Sus scrofa
0.0075
-
isocitrate pH 7.4, wild-type enzyme Sus scrofa
0.0092
-
isocitrate pH 7.4, mutant K212Y Sus scrofa
0.0098
-
isocitrate pH 7.4, mutant K212Q Sus scrofa
0.0099
-
NADP+ pH 7.4, wild-type enzyme Sus scrofa
0.014
-
NADP+ pH 7.4, mutant K212Q Sus scrofa
0.018
-
NADP+ pH 7.4, mutant K212R Sus scrofa
0.116
-
isocitrate pH 7.4, mutant K212R Sus scrofa
0.131
-
Mn2+ pH 7.4, mutant Y140K Sus scrofa
0.252
-
Mn2+ pH 7.4, mutant K212R Sus scrofa

Localization

Localization Comment Organism GeneOntology No. Textmining
mitochondrion
-
Sus scrofa 5739
-

Metals/Ions

Metals/Ions Comment Organism Structure
Mn2+
-
Sus scrofa

Molecular Weight [Da]

Molecular Weight [Da] Molecular Weight Maximum [Da] Comment Organism
43000
-
2 * 43000, recombinant wild-type and mutant enzymes, SDS-PAGE Sus scrofa
92000
-
recombinant wild-type and mutant enzymes, native PAGE Sus scrofa

Organism

Organism UniProt Comment Textmining
Sus scrofa P33198
-
-

Purification (Commentary)

Purification (Comment) Organism
recombinant wild-type and mutant enzymes from Escherichia coli, to homogeneity Sus scrofa

Reaction

Reaction Comment Organism Reaction ID
isocitrate + NADP+ = 2-oxoglutarate + CO2 + NADPH + H+ Tyr140 and Lys212 are required for catalytic activity, Tyr140 is the general acid that protonates the substrate after decarboxylation, Lys212 lowers as a positively charged residue the pK of the nearby ionizable group in the enzyme-substrate complex and stabilizes the carbanion formed initially on substrate decarboxylation Sus scrofa

Source Tissue

Source Tissue Comment Organism Textmining
heart
-
Sus scrofa
-

Specific Activity [micromol/min/mg]

Specific Activity Minimum [µmol/min/mg] Specific Activity Maximum [µmol/min/mg] Comment Organism
0.003
-
purified recombinant mutant K212Y, forward reaction Sus scrofa
0.065
-
purified recombinant mutant K212Q, forward reaction Sus scrofa
0.08
-
purified recombinant mutant Y140E, forward reaction Sus scrofa
0.083
-
purified recombinant mutant Y140T, forward reaction Sus scrofa
0.102
-
purified recombinant mutant Y140F, forward reaction Sus scrofa
0.179
-
purified recombinant mutant Y140K, forward reaction Sus scrofa
3.16
-
purified recombinant mutant K212R, forward reaction Sus scrofa
35.1
-
purified recombinant wild-type enzyme, forward reaction Sus scrofa

Substrates and Products (Substrate)

Substrates Comment Substrates Organism Products Comment (Products) Rev. Reac.
isocitrate + NADP+
-
Sus scrofa 2-oxoglutarate + CO2 + NADPH + H+
-
r

Subunits

Subunits Comment Organism
dimer 2 * 43000, recombinant wild-type and mutant enzymes, SDS-PAGE Sus scrofa

Synonyms

Synonyms Comment Organism
NADP-dependent isocitrate dehydrogenase
-
Sus scrofa

Temperature Optimum [°C]

Temperature Optimum [°C] Temperature Optimum Maximum [°C] Comment Organism
25
-
forward reaction, assay at Sus scrofa
37
-
reverse reaction, assay at Sus scrofa

pH Optimum

pH Optimum Minimum pH Optimum Maximum Comment Organism
7.4
-
assay at Sus scrofa

pH Range

pH Minimum pH Maximum Comment Organism
additional information
-
pH-profile, recombinant wild-type and mutant enzymes Sus scrofa

Cofactor

Cofactor Comment Organism Structure
NADP+
-
Sus scrofa
NADPH
-
Sus scrofa