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Literature summary for 1.5.1.20 extracted from

  • Trimmer, E.E.; Ballou, D.P.; Galloway, L.J.; Scannell, S.A.; Brinker, D.R.; Casas, K.R.
    Aspartate 120 of Escherichia coli methylenetetrahydrofolate reductase: evidence for major role in folate binding and catalysis and a minor role in flavin reactivity (2005), Biochemistry, 44, 6809-6822.
    View publication on PubMed
Search for more literature for 1.5.1.20

Cloned(Commentary)

Cloned (Comment) Organism
expression of wild-type and mutant enzymes in Escherichia coli strain AB109 and AB1909-(DE3)7D Escherichia coli

Protein Variants

Protein Variants Comment Organism
D120A site-directed mutagenesis, decreased catalytic efficiency in the folate oxidative half-reaction, loss of negative charge near the flavin, small increasing effects on the NADH reductive half reaction Escherichia coli
D120K site-directed mutagenesis, decreased catalytic efficiency in the folate oxidative half-reaction, loss of negative charge near the flavin, small increasing effects on the NADH reductive half reaction Escherichia coli
D120N site-directed mutagenesis, decreased catalytic efficiency in the folate oxidative half-reaction, loss of negative charge near the flavin, small increasing effects on the NADH reductive half reaction Escherichia coli
D120S site-directed mutagenesis, decreased catalytic efficiency in the folate oxidative half-reaction, loss of negative charge near the flavin, small increasing effects on the NADH reductive half reaction Escherichia coli
D120V site-directed mutagenesis, decreased catalytic efficiency in the folate oxidative half-reaction, loss of negative charge near the flavin, small increasing effects on the NADH reductive half reaction Escherichia coli

Inhibitors

Inhibitors Comment Organism Structure
methylenetetrahydrofolate
-
 Escherichia coli
NADH
-
 Escherichia coli

KM Value [mM]

KM Value [mM] KM Value Maximum [mM] Substrate Comment Organism Structure
additional information
-
 additional information redox potentials of wild-type and mutant enzymes, kinetic mechanism, and rapid-reaction kinetics for the half-reactions, steady-state kinetics Escherichia coli
0.0004
-
 methylenetetrahydrofolate recombinant wild-type enzyme, pH 7.2, 4°C Escherichia coli
0.0035
-
 NADH recombinant wild-type enzyme, pH 7.2, 4°C, the mutant enzymes all show a Km below 0.0035 mM Escherichia coli
0.017
-
 methylenetetrahydrofolate recombinant mutant D120N, pH 7.2, 4°C Escherichia coli
0.043
-
 methylenetetrahydrofolate recombinant mutant D120S, pH 7.2, 4°C Escherichia coli
0.099
-
 methylenetetrahydrofolate recombinant mutant D120A, pH 7.2, 4°C Escherichia coli
0.142
-
 methylenetetrahydrofolate recombinant mutant D120K, pH 7.2, 4°C Escherichia coli
0.187
-
 methylenetetrahydrofolate recombinant mutant D120V, pH 7.2, 4°C Escherichia coli

Natural Substrates/ Products (Substrates)

Natural Substrates Organism Comment (Nat. Sub.) Natural Products Comment (Nat. Pro.) Rev. Reac.
5-methyltetrahydrofolate + NAD+ Escherichia coli
-
 5,10-methylenetetrahydrofolate + NADH
-
 r

Organism

Organism UniProt Comment Textmining
Escherichia coli
-
-
-

Reaction

Reaction Comment Organism Reaction ID
5-methyltetrahydrofolate + NAD(P)+ = 5,10-methylenetetrahydrofolate + NAD(P)H + H+ substrate binding and catalyic mechanism involve Asp120, half-reaction mechanisms Escherichia coli
a

Substrates and Products (Substrate)

Substrates Comment Substrates Organism Products Comment (Products) Rev. Reac.
(6R,S)-5,10-methylenetetrahydrofolate + ?
-
 Escherichia coli ?
-
 ?
5-methyltetrahydrofolate + NAD+
-
 Escherichia coli 5,10-methylenetetrahydrofolate + NADH
-
 r

Synonyms

Synonyms Comment Organism
methylenetetrahydrofolate reductase
-
 Escherichia coli
MTHFR
-
 Escherichia coli

Turnover Number [1/s]

Turnover Number Minimum [1/s] Turnover Number Maximum [1/s] Substrate Comment Organism Structure
0.007
-
 methylenetetrahydrofolate recombinant mutant D120N, pH 7.2, 4°C Escherichia coli
0.011
-
 methylenetetrahydrofolate recombinant mutant D120K, pH 7.2, 4°C Escherichia coli
0.017
-
 methylenetetrahydrofolate recombinant mutant D120V, pH 7.2, 4°C Escherichia coli
0.02
-
 methylenetetrahydrofolate recombinant mutant D120S, pH 7.2, 4°C Escherichia coli
0.022
-
 methylenetetrahydrofolate recombinant mutant D120A, pH 7.2, 4°C Escherichia coli
2.2
-
 methylenetetrahydrofolate recombinant wild-type enzyme, pH 7.2, 4°C Escherichia coli

pH Optimum

pH Optimum Minimum pH Optimum Maximum Comment Organism
7.2
-
 assay at Escherichia coli

Cofactor

Cofactor Comment Organism Structure
FAD redox properties of enzyme-bound FAD are influenced by Asp120, which electrostatically stabilizes putative 5-iminium cation intermediate during catalysis Escherichia coli
NAD+
-
 Escherichia coli
NADH
-
 Escherichia coli

Ki Value [mM]

Ki Value [mM] Ki Value maximum [mM] Inhibitor Comment Organism Structure
0.014
-
 NADH recombinant wild-type enzyme, pH 7.2, 4°C, the mutant enzymes all show a Km below 0.0035 mM Escherichia coli
0.061
-
 methylenetetrahydrofolate recombinant wild-type enzyme, pH 7.2, 4°C Escherichia coli