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Literature summary extracted from

  • Kim, T.K.; Colman, R.F.
    Ser95, Asn97, and Thr78 are important for the catalytic function of porcine NADP-dependent isocitrate dehydrogenase (2005), Protein Sci., 14, 140-147.
    View publication on PubMedView publication on EuropePMC

Cloned(Commentary)

EC Number Cloned (Comment) Organism
1.1.1.42 expression of wild-type and mutant enzymes in Escherichia coli as maltose-binding fusion proteins Sus scrofa

Protein Variants

EC Number Protein Variants Comment Organism
1.1.1.42 N97A site-directed mutagenesis, decreased Vmax compared to the wild-type enzyme, slightly affected Km values, but increased pKa of the ionizable metal-liganded hydroxyl of enzyme-bound isocitrate compared to the wild-type enzyme Sus scrofa
1.1.1.42 N97D site-directed mutagenesis, highly decreased Vmax compared to the wild-type enzyme Sus scrofa
1.1.1.42 S95A site-directed mutagenesis, decreased Vmax, and increased Km for isocitrate and Mn2+ compared to the wild-type enzyme Sus scrofa
1.1.1.42 S95D site-directed mutagenesis, highly decreased Vmax compared to the wild-type enzyme Sus scrofa
1.1.1.42 T78A site-directed mutagenesis, decreased Vmax, and increased Km for isocitrate and Mn2+ compared to the wild-type enzyme Sus scrofa
1.1.1.42 T78D site-directed mutagenesis, decreased Vmax compared to the wild-type enzyme Sus scrofa

KM Value [mM]

EC Number KM Value [mM] KM Value Maximum [mM] Substrate Comment Organism Structure
1.1.1.42 0.00011
-
 Mn2+ recombinant wild-type enzyme, pH 7.4, 25°C Sus scrofa
1.1.1.42 0.0004
-
 Mn2+ recombinant mutant N97A, pH 7.4, 25°C Sus scrofa
1.1.1.42 0.0012
-
 NADP+ recombinant mutant N97D, pH 7.4, 25°C Sus scrofa
1.1.1.42 0.0024
-
 Mn2+ recombinant mutant N97D, pH 7.4, 25°C Sus scrofa
1.1.1.42 0.0033
-
 Mn2+ recombinant mutant T78A, pH 7.4, 25°C Sus scrofa
1.1.1.42 0.0049
-
 NADP+ recombinant mutant N97A, pH 7.4, 25°C Sus scrofa
1.1.1.42 0.0051
-
 NADP+ recombinant mutant S95A, pH 7.4 Sus scrofa
1.1.1.42 0.0053
-
 NADP+ recombinant mutant T78A, pH 7.4, 25°C Sus scrofa
1.1.1.42 0.0075
-
 isocitrate recombinant wild-type enzyme, pH 7.4, 25°C Sus scrofa
1.1.1.42 0.0099
-
 NADP+ recombinant wild-type enzyme, pH 7.4, 25°C Sus scrofa
1.1.1.42 0.0108
-
 NADP+ recombinant mutant T78D, pH 7.4, 25°C Sus scrofa
1.1.1.42 0.0112
-
 Mn2+ recombinant mutant S95A, pH 7.4, 25°C Sus scrofa
1.1.1.42 0.014
-
 isocitrate recombinant mutant N97A, pH 7.4, 25°C Sus scrofa
1.1.1.42 0.0237
-
 NADP+ recombinant mutant S95D, pH 7.4, 25°C Sus scrofa
1.1.1.42 0.0288
-
 Mn2+ recombinant mutant T78D, pH 7.4, 25°C Sus scrofa
1.1.1.42 0.066
-
 isocitrate recombinant mutant T78A, pH 7.4, 25°C Sus scrofa
1.1.1.42 0.0668
-
 Mn2+ recombinant mutant S95D, pH 7.4, 25°C Sus scrofa
1.1.1.42 0.072
-
 isocitrate recombinant mutant N97D, pH 7.4, 25°C Sus scrofa
1.1.1.42 0.101
-
 isocitrate recombinant mutant S95A, pH 7.4, 25°C Sus scrofa
1.1.1.42 0.367
-
 isocitrate recombinant mutant T78D, pH 7.4, 25°C Sus scrofa
1.1.1.42 2.07
-
 isocitrate recombinant mutant S95D, pH 7.4, 25°C Sus scrofa

Metals/Ions

EC Number Metals/Ions Comment Organism Structure
1.1.1.42 Mn2+ as Mn2+-isocitrate complex, Ser95, Asn97, and Thr78 are involved in binding Sus scrofa

Molecular Weight [Da]

EC Number Molecular Weight [Da] Molecular Weight Maximum [Da] Comment Organism
1.1.1.42 87000
-
 recombinant mutant N97A, native PAGE Sus scrofa
1.1.1.42 94000
-
 recombinant mutant T78A, native PAGE Sus scrofa
1.1.1.42 97000
-
 recombinant mutant N97D, native PAGE Sus scrofa
1.1.1.42 98000
-
 recombinant mutant T78D, native PAGE Sus scrofa
1.1.1.42 101000
-
 recombinant mutant S95A, native PAGE Sus scrofa
1.1.1.42 104000
-
 recombinant wild-type enzyme and mutant S95D, native PAGE Sus scrofa

Natural Substrates/ Products (Substrates)

EC Number Natural Substrates Organism Comment (Nat. Sub.) Natural Products Comment (Nat. Pro.) Rev. Reac.
1.1.1.42 isocitrate + NADP+ Sus scrofa
-
 2-oxoglutarate + CO2 + NADPH + H+
-
 ?

Organism

EC Number Organism UniProt Comment Textmining
1.1.1.42 Sus scrofa
-
-
-

Purification (Commentary)

EC Number Purification (Comment) Organism
1.1.1.42 recombinant wild-type and mutant enzymes from Escherichia coli, cleavage of the fusion proteins by thrombin, to homogeneity by amylose affinity chromatography Sus scrofa

Reaction

EC Number Reaction Comment Organism Reaction ID
1.1.1.42 isocitrate + NADP+ = 2-oxoglutarate + CO2 + NADPH + H+ Ser95, Asn97, and Thr78 are important for the catalysis having distinguishable functions, overview Sus scrofa
a

Specific Activity [micromol/min/mg]

EC Number Specific Activity Minimum [µmol/min/mg] Specific Activity Maximum [µmol/min/mg] Comment Organism
1.1.1.42 0.06
-
 purified recombinant mutant S95D Sus scrofa
1.1.1.42 0.44
-
 purified recombinant mutant N97D Sus scrofa
1.1.1.42 5.29
-
 purified recombinant mutant N97A Sus scrofa
1.1.1.42 6.22
-
 purified recombinant mutant T78D Sus scrofa
1.1.1.42 7.18
-
 purified recombinant mutant S95A Sus scrofa
1.1.1.42 12.7
-
 purified recombinant mutant T78A Sus scrofa
1.1.1.42 35.1
-
 purified recombinant wild-type enzyme Sus scrofa

Substrates and Products (Substrate)

EC Number Substrates Comment Substrates Organism Products Comment (Products) Rev. Reac.
1.1.1.42 isocitrate + NADP+
-
 Sus scrofa 2-oxoglutarate + CO2 + NADPH + H+
-
 ?
1.1.1.42 isocitrate + NADP+ Ser95, Asn97, and Thr78 are involved in substrate binding Sus scrofa 2-oxoglutarate + CO2 + NADPH + H+
-
 ?

Subunits

EC Number Subunits Comment Organism
1.1.1.42 dimer wild-type and mutant enzymes, circular dichroism and native PAGE Sus scrofa

Synonyms

EC Number Synonyms Comment Organism
1.1.1.42 NADP-dependent isocitrate dehydrogenase
-
 Sus scrofa

Temperature Optimum [°C]

EC Number Temperature Optimum [°C] Temperature Optimum Maximum [°C] Comment Organism
1.1.1.42 25
-
 assay at Sus scrofa

pH Optimum

EC Number pH Optimum Minimum pH Optimum Maximum Comment Organism
1.1.1.42 7.4
-
 assay at Sus scrofa

pH Range

EC Number pH Minimum pH Maximum Comment Organism
1.1.1.42 5 8 pH-profile of wild-type and alanine-substituted mutant enzymes Sus scrofa

Cofactor

EC Number Cofactor Comment Organism Structure
1.1.1.42 NADP+
-
 Sus scrofa