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Literature summary for 1.1.1.40 extracted from

  • Detarsio, E.; Wheeler, M.C.; Campos Bermudez, V.A.; Andreo, C.S.; Drincovich, M.F.
    Maize C4 NADP-malic enzyme. Expression in Escherichia coli and characterization of site-directed mutants at the putative nucleoside-binding sites (2003), J. Biol. Chem., 278, 13757-13764.
    View publication on PubMed

Cloned(Commentary)

Cloned (Comment) Organism
overexpression of His-tagged wild-type and mutants in Escherichia coli strain DH5alpha, functional complementation of the enzyme deficient Escherichia coli triple mutant strain EJ1321 Zea mays

Protein Variants

Protein Variants Comment Organism
A387G site directed mutagenesis, mutation at the NADP+ binding site, mutant shows 48fold decreased kcat and 4.3 and 5.8fold increased Km for NADP+ and L-malate, respectively, compared to the wild-type enzyme, no activity with NAD+ Zea mays
A392G site directed mutagenesis, mutation at the NADP+ binding site, mutant shows unaltered kcat, but 3.5 and 2.6fold increased Km for NADP+ and L-malate, respectively, and increased activity with NAD+ compared to the wild-type enzyme Zea mays
additional information construction of the Gsite5V and Gsite2V mutants, mutation at the NADP+ binding site Zea mays
R237L site directed mutagenesis, mutation at the NADP+ binding site, mutant shows 530fold decreased kcat and 36.3 and 15.3fold increased Km for NADP+ and L-malate, respectively, compared to the wild-type enzyme, no activity with NAD+ Zea mays

Inhibitors

Inhibitors Comment Organism Structure
L-malate pH 7.0, high concentration Zea mays

KM Value [mM]

KM Value [mM] KM Value Maximum [mM] Substrate Comment Organism Structure
0.008
-
NADP+ wild-type enzyme, pH 7.0 Zea mays
0.0086
-
NADP+ recombinant enzyme, pH 7.0 Zea mays
0.03
-
NADP+ mutant A392G, pH 7.0 Zea mays
0.037
-
NADP+ mutant A387G, pH 7.0 Zea mays
0.19
-
L-malate recombinant enzyme, pH 7.0 Zea mays
0.23
-
L-malate wild-type enzyme, pH 7.0 Zea mays
0.29
-
NADP+ mutant R237L, pH 7.0 Zea mays
0.5
-
L-malate mutant A392G, pH 7.0 Zea mays
1.1
-
L-malate mutant A387G, pH 7.0 Zea mays
2.9
-
L-malate mutant R237L, pH 7.0 Zea mays
6
-
NAD+ mutant A392G, pH 7.0 Zea mays
8.1
-
NAD+ wild-type enzyme, pH 7.0 Zea mays

Localization

Localization Comment Organism GeneOntology No. Textmining
chloroplast
-
Zea mays 9507
-

Metals/Ions

Metals/Ions Comment Organism Structure
Mg2+
-
Zea mays

Molecular Weight [Da]

Molecular Weight [Da] Molecular Weight Maximum [Da] Comment Organism
62000
-
x * 62000, detagged recombinant wild-type enzyme Zea mays
110000
-
recombinant dimeric enzyme, gel filtration at pH 7.0, minor peak Zea mays
226000
-
recombinant tetrameric enzyme, gel filtration at pH 8.0, major peak Zea mays

Natural Substrates/ Products (Substrates)

Natural Substrates Organism Comment (Nat. Sub.) Natural Products Comment (Nat. Pro.) Rev. Reac.
(S)-malate + NAD(P)+ Zea mays the unique and specialized C4-type enzyme has evolved fro the C3-type enzyme pyruvate + CO2 + NAD(P)H
-
?

Organism

Organism UniProt Comment Textmining
Zea mays P16243
-
-

Purification (Commentary)

Purification (Comment) Organism
recombinant His-tagged mutant enzyme to homogeneity from Escherichia coli by metal affinity chromatography Zea mays

Source Tissue

Source Tissue Comment Organism Textmining
leaf
-
Zea mays
-

Substrates and Products (Substrate)

Substrates Comment Substrates Organism Products Comment (Products) Rev. Reac.
(S)-malate + NAD(P)+
-
Zea mays pyruvate + CO2 + NAD(P)H
-
?
(S)-malate + NAD(P)+ the unique and specialized C4-type enzyme has evolved fro the C3-type enzyme Zea mays pyruvate + CO2 + NAD(P)H
-
?
(S)-malate + NAD+
-
Zea mays pyruvate + CO2 + NADH
-
?

Subunits

Subunits Comment Organism
More extent of oligomerization is pH-dependent, 3D-structure analysis Zea mays
oligomer x * 62000, detagged recombinant wild-type enzyme Zea mays

Synonyms

Synonyms Comment Organism
NADP-malic enzyme
-
Zea mays
NADP-ME
-
Zea mays

Turnover Number [1/s]

Turnover Number Minimum [1/s] Turnover Number Maximum [1/s] Substrate Comment Organism Structure
0.38
-
NADP+ mutant R237L, pH 7.0 Zea mays
4.2
-
NADP+ mutant A387G, pH 7.0 Zea mays
13.5
-
NAD+ wild-type enzyme, pH 7.0 Zea mays
30.8
-
NADP+ recombinant enzyme, pH 7.0 Zea mays
40.6
-
NAD+ mutant A392G, pH 7.0 Zea mays
200.3
-
NADP+ mutant A392G, pH 7.0 Zea mays
201.3
-
NADP+ wild-type enzyme, pH 7.0 Zea mays

pH Optimum

pH Optimum Minimum pH Optimum Maximum Comment Organism
8
-
recombinant enzyme Zea mays

Cofactor

Cofactor Comment Organism Structure
NAD+ the recombinant wild-type enzyme shows a low intrinsic activity with NAD+, best at pH 7.0, mutant A392G shows increased activity with NAD+ compared to the wild-type enzyme Zea mays
NADP+ binding site structure, Arg237 is important for cofactor binding and specificity Zea mays

Ki Value [mM]

Ki Value [mM] Ki Value maximum [mM] Inhibitor Comment Organism Structure
4.7
-
L-malate recombinant enzyme, pH 7.0 Zea mays

pI Value

Organism Comment pI Value Maximum pI Value
Zea mays recombinant enzyme, isoelectric focusing
-
7.1