Cloned (Comment) | Organism |
---|---|
expression of wild-type and mutant enzymes in Escherichia coli strain BL21(DE3) | Zea mays |
Protein Variants | Comment | Organism |
---|---|---|
C192A | site-directed mutagenesis of isozyme ZmC4-NADP-ME, the mutation does not affect the tetrameric state, the mutant displays lower malate affinity than the wild-type enzyme | Zea mays |
C192A | marked decrease in kcat value, less than 10% of wild-type, with concomitant increase in Km value for NADP+. Unlike wild-type, activity is not significantly changed in presence of oxidant iodosobenzoate | Zea mays |
C231A | site-directed mutagenesis of isozyme ZmC4-NADP-ME, the mutation does not affect the tetrameric state, the mutant displays lower malate affinity than the wild-type enzyme | Zea mays |
C231A | marked decrease in kcat value, less than 10% of wild-type, with concomitant increase in Km value for NADP+. Similar to wild-type, activity decreases in presence of oxidant iodosobenzoate | Zea mays |
C246A | site-directed mutagenesis of isozyme ZmC4-NADP-ME, the mutation does not affect the tetrameric state, C246A exhibits a nearly 5fold increase in its affinity towards NADP+ and a 3fold decrease for malate compared to the wild-tpe enzyme | Zea mays |
C246A | marked decrease in kcat value, less than 10% of wild-type, with concomitant increase in Km value for NADP+. Unlike wild-type, activity is not significantly changed in presence of oxidant iodosobenzoate | Zea mays |
C270A | site-directed mutagenesis of isozyme ZmC4-NADP-ME, the mutation does not affect the tetrameric state, the mutant displays lower malate affinity than the wild-type enzyme | Zea mays |
C270A | marked decrease in kcat value, less than 10% of wild-type, with concomitant increase in Km value for NADP+. Unlike wild-type, activity is not significantly changed in presence of oxidant iodosobenzoate | Zea mays |
Inhibitors | Comment | Organism | Structure |
---|---|---|---|
CuCl2 | about 40% loss of activity within 60 min | Zea mays | |
Diamide | 2 mM, time-dependent decrease in activity reaching about 40% of initial activity after 60 min. In presence of dithiothreitol, a complete recovery is observed after 90 min. Enzyme oxidation decreases the catalytic activity. No severe loss of protein secondary structure takes place after oxidation; about 40% loss of activity within 60 min | Zea mays | |
Iodosobenzoate | 1 mM, time-dependent decrease in activity reaching about 40% of initial activity after 60 min; about 40% loss of activity within 60 min | Zea mays | |
additional information | ZmnonC4-NADP-ME activity is not significantly modified by any chemical oxidant in 60 min | Zea mays |
KM Value [mM] | KM Value Maximum [mM] | Substrate | Comment | Organism | Structure |
---|---|---|---|---|---|
0.00014 | - |
(S)-malate | pH 8.0, 30°C, mutant C246A | Zea mays | |
0.00015 | - |
(S)-malate | pH 7.0, 30°C, wild-type, treated with dithiothreitol | Zea mays | |
0.00017 | - |
(S)-malate | pH 8.0, 30°C, wild-type, treated with diamide | Zea mays | |
0.00039 | - |
(S)-malate | pH 8.0, 30°C, wild-type, treated with dithiothreitol | Zea mays | |
0.0008 | - |
(S)-malate | pH 7.0, 30°C, wild-type, treated with diamide | Zea mays | |
0.0017 | - |
(S)-malate | pH 8.0, 30°C, mutant C231A | Zea mays | |
0.003 | - |
(S)-malate | pH 8.0, 30°C, mutant C192A | Zea mays | |
0.0036 | - |
NADP+ | pH 7.0, 30°C, oxidized isozyme ZmC4-NADP-ME | Zea mays | |
0.0036 | - |
NADP+ | pH 7.0, 30°C, wild-type, treated with diamide | Zea mays | |
0.0043 | - |
(S)-malate | pH 8.0, 30°C, mutant C270A | Zea mays | |
0.0053 | - |
NADP+ | pH 8.0, 30°C, reduced isozyme ZmC4-NADP-ME, mutant C246A | Zea mays | |
0.0053 | - |
NADP+ | pH 8.0, 30°C, mutant C246A | Zea mays | |
0.0088 | - |
NADP+ | pH 8.0, 30°C, oxidized isozyme ZmC4-NADP-ME | Zea mays | |
0.0088 | - |
NADP+ | pH 8.0, 30°C, wild-type, treated with diamide | Zea mays | |
0.0135 | - |
NADP+ | pH 8.0, 30°C, reduced isozyme ZmC4-NADP-ME, mutant C270A | Zea mays | |
0.0135 | - |
NADP+ | pH 8.0, 30°C, mutant C270A | Zea mays | |
0.0147 | - |
NADP+ | pH 7.0, 30°C, reduced isozyme ZmC4-NADP-ME | Zea mays | |
0.0147 | - |
NADP+ | pH 7.0, 30°C, wild-type, treated with dithiothreitol | Zea mays | |
0.015 | - |
NADP+ | pH 8.0, 30°C, reduced isozyme ZmC4-NADP-ME, mutant C231A | Zea mays | |
0.015 | - |
NADP+ | pH 8.0, 30°C, mutant C231A | Zea mays | |
0.0154 | - |
NADP+ | pH 8.0, 30°C, reduced isozyme ZmC4-NADP-ME, mutant C192A | Zea mays | |
0.0154 | - |
NADP+ | pH 8.0, 30°C, mutant C192A | Zea mays | |
0.0276 | - |
NADP+ | pH 8.0, 30°C, reduced isozyme ZmC4-NADP-ME | Zea mays | |
0.0276 | - |
NADP+ | pH 8.0, 30°C, wild-type, treated with dithiothreitol | Zea mays | |
0.08 | - |
(S)-malate | pH 7.0, 30°C, oxidized isozyme ZmC4-NADP-ME | Zea mays | |
0.14 | - |
(S)-malate | pH 8.0, 30°C, reduced isozyme ZmC4-NADP-ME, mutant C246A | Zea mays | |
0.15 | - |
(S)-malate | pH 7.0, 30°C, reduced isozyme ZmC4-NADP-ME | Zea mays | |
0.17 | - |
(S)-malate | pH 8.0, 30°C, oxidized isozyme ZmC4-NADP-ME | Zea mays | |
0.39 | - |
(S)-malate | pH 8.0, 30°C, reduced isozyme ZmC4-NADP-ME | Zea mays | |
1.7 | - |
(S)-malate | pH 8.0, 30°C, reduced isozyme ZmC4-NADP-ME, mutant C231A | Zea mays | |
3 | - |
(S)-malate | pH 8.0, 30°C, reduced isozyme ZmC4-NADP-ME, mutant C192A | Zea mays | |
4.3 | - |
(S)-malate | pH 8.0, 30°C, reduced isozyme ZmC4-NADP-ME, mutant C270A | Zea mays |
Localization | Comment | Organism | GeneOntology No. | Textmining |
---|---|---|---|---|
chloroplast | isozymes ZmC4-NADP-ME and ZmnonC4-NADP-ME | Zea mays | 9507 | - |
plastid | - |
Zea mays | 9536 | - |
Metals/Ions | Comment | Organism | Structure |
---|---|---|---|
Mg2+ | required | Zea mays |
Natural Substrates | Organism | Comment (Nat. Sub.) | Natural Products | Comment (Nat. Pro.) | Rev. | Reac. |
---|---|---|---|---|---|---|
(S)-malate + NADP+ | Zea mays | - |
pyruvate + CO2 + NADPH | - |
? |
Organism | UniProt | Comment | Textmining |
---|---|---|---|
Zea mays | - |
isozymes ZmC4-NADP-ME and ZmnonC4-NADP-ME | - |
Zea mays | P16243 | - |
- |
Purification (Comment) | Organism |
---|---|
recombinant wild-type and mutant enzymes from Escherichia coli strain BL21(DE3) to homogeneity | Zea mays |
Renatured (Comment) | Organism |
---|---|
presence of 2 mM diamide leads to a time-dependent decrease in activity, reaching about 40% of initial activity after 60 min. After addition of dithiothreitol, a complete recovery is observed after 90 min | Zea mays |
Source Tissue | Comment | Organism | Textmining |
---|---|---|---|
bundle sheath cell | isozyme ZmC4-NADP-ME | Zea mays | - |
Substrates | Comment Substrates | Organism | Products | Comment (Products) | Rev. | Reac. |
---|---|---|---|---|---|---|
(S)-malate + NADP+ | - |
Zea mays | pyruvate + CO2 + NADPH | - |
? |
Subunits | Comment | Organism |
---|---|---|
tetramer | recombinant ZmC4-NADP-ME wild-type and mutants | Zea mays |
Synonyms | Comment | Organism |
---|---|---|
C4-NADP-ME | isoform involved in C4 photosynthesis | Zea mays |
NADP-malic enzyme | - |
Zea mays |
NADP-ME | - |
Zea mays |
Temperature Optimum [°C] | Temperature Optimum Maximum [°C] | Comment | Organism |
---|---|---|---|
30 | - |
assay at | Zea mays |
Turnover Number Minimum [1/s] | Turnover Number Maximum [1/s] | Substrate | Comment | Organism | Structure |
---|---|---|---|---|---|
1.1 | - |
NADP+ | pH 8.0, 30°C, reduced isozyme ZmC4-NADP-ME, mutant C246A | Zea mays | |
1.1 | - |
(S)-malate | pH 8.0, 30°C, reduced isozyme ZmC4-NADP-ME, mutant C246A | Zea mays | |
1.1 | - |
NADP+ | pH 8.0, 30°C, mutant C246A | Zea mays | |
1.6 | - |
NADP+ | pH 8.0, 30°C, reduced isozyme ZmC4-NADP-ME, mutant C270A | Zea mays | |
1.6 | - |
(S)-malate | pH 8.0, 30°C, reduced isozyme ZmC4-NADP-ME, mutant C270A | Zea mays | |
1.6 | - |
NADP+ | pH 8.0, 30°C, mutant C270A | Zea mays | |
3.55 | - |
NADP+ | pH 7.0, 30°C, oxidized isozyme ZmC4-NADP-ME | Zea mays | |
3.55 | - |
(S)-malate | pH 7.0, 30°C, oxidized isozyme ZmC4-NADP-ME | Zea mays | |
3.55 | - |
NADP+ | pH 7.0, 30°C, wild-type, treated with diamide | Zea mays | |
5.6 | - |
NADP+ | pH 8.0, 30°C, reduced isozyme ZmC4-NADP-ME, mutant C192A | Zea mays | |
5.6 | - |
(S)-malate | pH 8.0, 30°C, reduced isozyme ZmC4-NADP-ME, mutant C192A | Zea mays | |
5.6 | - |
NADP+ | pH 8.0, 30°C, mutant C192A | Zea mays | |
8.35 | - |
NADP+ | pH 7.0, 30°C, reduced isozyme ZmC4-NADP-ME | Zea mays | |
8.35 | - |
(S)-malate | pH 7.0, 30°C, reduced isozyme ZmC4-NADP-ME | Zea mays | |
8.35 | - |
NADP+ | pH 7.0, 30°C, wild-type, treated with dithiothreitol | Zea mays | |
23.2 | - |
NADP+ | pH 8.0, 30°C, reduced isozyme ZmC4-NADP-ME, mutant C231A | Zea mays | |
23.2 | - |
(S)-malate | pH 8.0, 30°C, reduced isozyme ZmC4-NADP-ME, mutant C231A | Zea mays | |
23.2 | - |
NADP+ | pH 8.0, 30°C, mutant C231A | Zea mays | |
91 | - |
NADP+ | pH 8.0, 30°C, oxidized isozyme ZmC4-NADP-ME | Zea mays | |
91 | - |
(S)-malate | pH 8.0, 30°C, oxidized isozyme ZmC4-NADP-ME | Zea mays | |
91 | - |
NADP+ | pH 8.0, 30°C, wild-type, treated with diamide | Zea mays | |
269 | - |
NADP+ | pH 8.0, 30°C, reduced isozyme ZmC4-NADP-ME | Zea mays | |
269 | - |
(S)-malate | pH 8.0, 30°C, reduced isozyme ZmC4-NADP-ME | Zea mays | |
269 | - |
NADP+ | pH 8.0, 30°C, wild-type, treated with dithiothreitol | Zea mays |
pH Optimum Minimum | pH Optimum Maximum | Comment | Organism |
---|---|---|---|
7 | 8 | assay at | Zea mays |
Cofactor | Comment | Organism | Structure |
---|---|---|---|
NADP+ | - |
Zea mays |
General Information | Comment | Organism |
---|---|---|
metabolism | isoform C4-NADP-ME involved in C4 photosynthesis is modulated by redox status, and its oxidation produces a conformational change limiting the catalytic process, although inducing higher affinity binding of the substrates. Residues Cys192, Cys246, Cys270 and Cys410 are directly or indirectly implicated in C4-NADP-ME redox modulation | Zea mays |
additional information | reversal of ZmC4-NADP-ME oxidation by chemical reductants, e.g. iodosobenzoate and CuCl2, doe to the presence of thiol groups able to form disulfide bonds. Residues Cys192, Cys246, Cys270, and Cys410 may be directly or indirectly implicated in ZmC4-NADP-ME redox modulation. Redox regulation plays a key role in many plastid functions. Isozyme specific redox regulation of ZmC4-NADP-ME activity, the modulation is not observed in the case of isozyme ZmnonC4-NADP-ME. The replacement of Cys246 with serine in ZmnonC4-NADP-ME may be responsible for the absence of redox modulation | Zea mays |
physiological function | one isozyme exclusively expressed in the bundle sheath cells and involved in C4 photosynthesis, i.e. ZmC4-NADP-ME, and the other, ZmnonC4-NADP-ME, with housekeeping roles | Zea mays |
kcat/KM Value [1/mMs-1] | kcat/KM Value Maximum [1/mMs-1] | Substrate | Comment | Organism | Structure |
---|---|---|---|---|---|
0.12 | - |
NADP+ | pH 8.0, 30°C, mutant C270A | Zea mays | |
0.14 | - |
(S)-malate | pH 8.0, 30°C, reduced isozyme ZmC4-NADP-ME, mutant C246A | Zea mays | |
0.21 | - |
NADP+ | pH 8.0, 30°C, mutant C246A | Zea mays | |
0.36 | - |
NADP+ | pH 8.0, 30°C, mutant C192A | Zea mays | |
0.37 | - |
NADP+ | pH 8.0, 30°C, reduced isozyme ZmC4-NADP-ME, mutant C270A | Zea mays | |
0.37 | - |
(S)-malate | pH 8.0, 30°C, mutant C270A | Zea mays | |
1.5 | - |
NADP+ | pH 8.0, 30°C, mutant C231A | Zea mays | |
1.7 | - |
(S)-malate | pH 8.0, 30°C, reduced isozyme ZmC4-NADP-ME, mutant C231A | Zea mays | |
1.86 | - |
NADP+ | pH 8.0, 30°C, reduced isozyme ZmC4-NADP-ME, mutant C192A | Zea mays | |
1.9 | - |
(S)-malate | pH 8.0, 30°C, mutant C192A | Zea mays | |
3 | - |
(S)-malate | pH 8.0, 30°C, reduced isozyme ZmC4-NADP-ME, mutant C192A | Zea mays | |
4.3 | - |
(S)-malate | pH 8.0, 30°C, reduced isozyme ZmC4-NADP-ME, mutant C270A | Zea mays | |
7.8 | - |
(S)-malate | pH 8.0, 30°C, mutant C246A | Zea mays | |
7.86 | - |
NADP+ | pH 8.0, 30°C, reduced isozyme ZmC4-NADP-ME, mutant C246A | Zea mays | |
9.7 | - |
NADP+ | pH 8.0, 30°C, wild-type, treated with dithiothreitol | Zea mays | |
13.6 | - |
(S)-malate | pH 8.0, 30°C, mutant C231A | Zea mays | |
13.64 | - |
NADP+ | pH 8.0, 30°C, reduced isozyme ZmC4-NADP-ME, mutant C231A | Zea mays | |
689.7 | - |
(S)-malate | pH 8.0, 30°C, reduced isozyme ZmC4-NADP-ME | Zea mays | |
690 | - |
(S)-malate | pH 8.0, 30°C, wild-type, treated with dithiothreitol | Zea mays | |
9746 | - |
NADP+ | pH 8.0, 30°C, reduced isozyme ZmC4-NADP-ME | Zea mays |