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Literature summary extracted from

  • Clark, J.E.; Ljungdahl, L.G.
    Purification and properties of 5,10-methylenetetrahydrofolate reductase, an iron-sulfur flavoprotein from Clostridium formicoaceticum (1984), J. Biol. Chem., 259, 10845-10849.
    View publication on PubMed

General Stability

EC Number General Stability Organism
1.5.1.20 0.005 mM FAD stabilizes enzyme during purification procedure Clostridium formicaceticum

KM Value [mM]

EC Number KM Value [mM] KM Value Maximum [mM] Substrate Comment Organism Structure
1.5.1.20 0.12
-
 5-methyltetrahydrofolate
-
 Clostridium formicaceticum
1.5.1.20 11.1
-
 benzyl viologen
-
 Clostridium formicaceticum
1.5.7.1 0.01
-
 5-methyltetrahydrofolate
-
 Clostridium formicaceticum
1.5.7.1 11.1
-
 benzyl viologen
-
 Clostridium formicaceticum

Metals/Ions

EC Number Metals/Ions Comment Organism Structure
1.5.1.20 Iron 15.2 molecules iron per enzyme molecule, enzyme contains iron-sulfur clusters Clostridium formicaceticum
1.5.1.20 iron-sulfur centre enzyme contains 15.2 molecules iron and 19.5 molecules acid-labile sulfur as iron-sulfur clusters Clostridium formicaceticum
1.5.1.20 Zinc 2.3 molecules zinc per enzyme molecule Clostridium formicaceticum
1.5.7.1 Iron iron-sulfur protein, contains 15.2 iron and 19.5 acid labile sulfide per 237000 Da enzyme Clostridium formicaceticum
1.5.7.1 Zinc enzyme contains 2.3 zinc per 237000 Da enzyme Clostridium formicaceticum

Molecular Weight [Da]

EC Number Molecular Weight [Da] Molecular Weight Maximum [Da] Comment Organism
1.5.1.20 26000
-
 alpha4,beta4, 4 * 26000 + 4 * 35000, SDS-PAGE Clostridium formicaceticum
1.5.1.20 26000
-
 alpha2,beta2, 2 * 26000 + 2 * 35000, determined at pH 9, SDS-PAGE Clostridium formicaceticum
1.5.1.20 35000
-
 alpha4,beta4, 4 * 26000 + 4 * 35000, SDS-PAGE Clostridium formicaceticum
1.5.1.20 35000
-
 alpha2,beta2, 2 * 26000 + 2 * 35000, determined at pH 9, SDS-PAGE Clostridium formicaceticum
1.5.1.20 124000
-
 determination at pH 9, alpha2,beta2 enzyme form, PAGE Clostridium formicaceticum
1.5.1.20 237000
-
 gel filtration Clostridium formicaceticum
1.5.1.20 237000
-
 determination at pH 7.4, alpha4,beta4 enzyme form Clostridium formicaceticum
1.5.7.1 26000
-
 alpha4beta4, 4 * 26000 + 4 * 35000, SDS-PAGE Clostridium formicaceticum
1.5.7.1 35000
-
 alpha4beta4, 4 * 26000 + 4 * 35000, SDS-PAGE Clostridium formicaceticum
1.5.7.1 237000
-
 gel filtration Clostridium formicaceticum

Natural Substrates/ Products (Substrates)

EC Number Natural Substrates Organism Comment (Nat. Sub.) Natural Products Comment (Nat. Pro.) Rev. Reac.
1.5.1.20 5,10-methylenetetrahydrofolate + FADH2 Clostridium formicaceticum enzyme in pathway of synthesis of acetate from CO2 via formate and a series of reactions involving tetrahydrofolate and a corrinoid, FADH2 and reduced ferredoxin may serve as natural reductants for 5,10-methylenetetrahydrofolate 5-methyltetrahydrofolate + FAD
-
 ?
1.5.1.20 5,10-methylenetetrahydrofolate + reduced ferredoxin Clostridium formicaceticum enzyme in pathway of synthesis of acetate from CO2 via formate and a series of reactions involving tetrahydrofolate and a corrinoid, FADH2 and reduced ferredoxin may serve as natural reductants for 5,10-methylenetetrahydrofolate, electron transfer from reduced ferredoxin via iron-sulfur centers via enzyme-bound FAD to 5,10-methylenetetrahydrofolate 5-methyltetrahydrofolate + oxidized ferredoxin
-
 ?
1.5.1.20 5,10-methylenetetrahydrofolate + reduced ferredoxin Clostridium formicaceticum physiological important reaction: enzyme catalyzes reduction of 5,10-methylenetetrahydrofolate with reduced ferredoxin 5-methyltetrahydrofolate + oxidized ferredoxin
-
 ?

Organism

EC Number Organism UniProt Comment Textmining
1.5.1.20 Clostridium formicaceticum
-
 strain ATCC 23439
-
1.5.7.1 Clostridium formicaceticum
-
-
-

Oxidation Stability

EC Number Oxidation Stability Organism
1.5.1.20 oxygen labile, half-life: less than 1 h in aerobic buffer, sodium dithionite prevents inactivation by oxygen Clostridium formicaceticum
1.5.7.1 the enzyme loses 50% of the activity within 2 h in aerobic buffer (0.1 M Tris/HCl buffer, pH 7.4) with or without a reducing agent and in anaerobic buffer without a reducing agent Clostridium formicaceticum

Purification (Commentary)

EC Number Purification (Comment) Organism
1.5.1.20
-
 Clostridium formicaceticum
1.5.7.1
-
 Clostridium formicaceticum

Specific Activity [micromol/min/mg]

EC Number Specific Activity Minimum [µmol/min/mg] Specific Activity Maximum [µmol/min/mg] Comment Organism
1.5.1.20 139
-
-
 Clostridium formicaceticum
1.5.7.1 139
-
-
 Clostridium formicaceticum

Storage Stability

EC Number Storage Stability Organism
1.5.1.20 10°C, 50 mM Tris-HCl buffer, pH 7.4, 20% glycerol, 2 mM dithionite, in an anaerobic chamber, a few weeks, stable Clostridium formicaceticum
1.5.1.20 frozen, 50 mM Tris-HCl buffer, pH 7.4, 20% glycerol, 2 mM dithionite, longer time, 10-20% loss of activity Clostridium formicaceticum
1.5.7.1 10°C, pH 7.4, 50 mM Tris/HCl containing 20% glycerol and 2 mM dithionite in an anaerobic chamber Clostridium formicaceticum

Substrates and Products (Substrate)

EC Number Substrates Comment Substrates Organism Products Comment (Products) Rev. Reac.
1.5.1.20 5,10-methylenetetrahydrofolate + FADH2 enzyme in pathway of synthesis of acetate from CO2 via formate and a series of reactions involving tetrahydrofolate and a corrinoid, FADH2 and reduced ferredoxin may serve as natural reductants for 5,10-methylenetetrahydrofolate Clostridium formicaceticum 5-methyltetrahydrofolate + FAD
-
 ?
1.5.1.20 5,10-methylenetetrahydrofolate + reduced acceptor forward reaction: reduced acceptor is FADH2 Clostridium formicaceticum 5-methyltetrahydrofolate + oxidized acceptor
-
 r
1.5.1.20 5,10-methylenetetrahydrofolate + reduced acceptor reverse reaction: menadione as electron acceptor Clostridium formicaceticum 5-methyltetrahydrofolate + oxidized acceptor
-
 r
1.5.1.20 5,10-methylenetetrahydrofolate + reduced acceptor reverse reaction: FAD, rubredoxin, benzyl viologen and methylene blue as electron acceptor Clostridium formicaceticum 5-methyltetrahydrofolate + oxidized acceptor
-
 r
1.5.1.20 5,10-methylenetetrahydrofolate + reduced acceptor forward reaction: reduced ferredoxin as reduced acceptor Clostridium formicaceticum 5-methyltetrahydrofolate + oxidized acceptor
-
 r
1.5.1.20 5,10-methylenetetrahydrofolate + reduced ferredoxin enzyme in pathway of synthesis of acetate from CO2 via formate and a series of reactions involving tetrahydrofolate and a corrinoid, FADH2 and reduced ferredoxin may serve as natural reductants for 5,10-methylenetetrahydrofolate, electron transfer from reduced ferredoxin via iron-sulfur centers via enzyme-bound FAD to 5,10-methylenetetrahydrofolate Clostridium formicaceticum 5-methyltetrahydrofolate + oxidized ferredoxin
-
 ?
1.5.1.20 5,10-methylenetetrahydrofolate + reduced ferredoxin physiological important reaction: enzyme catalyzes reduction of 5,10-methylenetetrahydrofolate with reduced ferredoxin Clostridium formicaceticum 5-methyltetrahydrofolate + oxidized ferredoxin
-
 ?
1.5.1.20 5-methyltetrahydrofolate + NAD+
-
 Clostridium formicaceticum 5,10-methylenetetrahydrofolate + NADH + H+
-
 r
1.5.1.20 additional information no direct activity with pyridine nucleotides Clostridium formicaceticum ?
-
 ?
1.5.1.20 nitrate + reduced benzyl viologen
-
 Clostridium formicaceticum nitrite + benzyl viologen
-
 ?
1.5.7.1 5,10-methylenetetrahydrofolate + FADH2
-
 Clostridium formicaceticum 5,10-methylenetetrahydrofolate + FAD
-
 ?
1.5.7.1 5,10-methylenetetrahydrofolate + reduced ferredoxin
-
 Clostridium formicaceticum 5-methyltetrahydrofolate + ferredoxin
-
 ?
1.5.7.1 5-methyltetrahydrofolate + benzyl viologen
-
 Clostridium formicaceticum 5,10-methylenetetrahydrofolate + reduced benzyl viologen
-
 ?
1.5.7.1 5-methyltetrahydrofolate + benzyl viologen 11% of the activity with methylene blue Clostridium formicaceticum 5,10-methylenetetrahydrofolate + reduced benzyl viologen
-
 ?
1.5.7.1 5-methyltetrahydrofolate + FAD 11% of the activity with methylene blue Clostridium formicaceticum 5,10-methylenetetrahydrofolate + FADH2
-
 ?
1.5.7.1 5-methyltetrahydrofolate + menadione 25% of the activity with methylene blue Clostridium formicaceticum 5,10-methylenetetrahydrofolate + reduced menadiol
-
 ?
1.5.7.1 5-methyltetrahydrofolate + rubredoxin 11% of the activity with methylene blue Clostridium formicaceticum 5,10-methylenetetrahydrofolate + reduced rubredoxin
-
 ?
1.5.7.1 additional information no activity with pyridine nucleotides Clostridium formicaceticum ?
-
 ?

Subunits

EC Number Subunits Comment Organism
1.5.1.20 octamer alpha4,beta4, 4 * 26000 + 4 * 35000, SDS-PAGE Clostridium formicaceticum
1.5.1.20 octamer determined at pH 7.4 Clostridium formicaceticum
1.5.1.20 tetramer alpha2,beta2, 2 * 26000 + 2 * 35000, determined at pH 9, SDS-PAGE Clostridium formicaceticum
1.5.7.1 octamer alpha4beta4, 4 * 26000 + 4 * 35000, SDS-PAGE Clostridium formicaceticum

Synonyms

EC Number Synonyms Comment Organism
1.5.7.1 5,10-methylenetetrahydrofolate reductase
-
 Clostridium formicaceticum

Temperature Optimum [°C]

EC Number Temperature Optimum [°C] Temperature Optimum Maximum [°C] Comment Organism
1.5.1.20 37
-
 assay at Clostridium formicaceticum

pH Optimum

EC Number pH Optimum Minimum pH Optimum Maximum Comment Organism
1.5.1.20 7.8 8
-
 Clostridium formicaceticum
1.5.7.1 7.8 8
-
 Clostridium formicaceticum

pH Stability

EC Number pH Stability pH Stability Maximum Comment Organism
1.5.1.20 7.1 7.6 at this pH-range enzyme is more stable in Tris-HCl than in triethanolamine-HCl or phosphate buffers Clostridium formicaceticum
1.5.7.1 7.1 7.6 the enzyme is more stable in Tris/HCl than in triethanolamine/HCl or phosphate buffer Clostridium formicaceticum

Cofactor

EC Number Cofactor Comment Organism Structure
1.5.1.20 FAD 1.7 molecules enzyme-bound FAD per enzyme molecule Clostridium formicaceticum
1.5.1.20 FAD flavoprotein, oxidized acceptor in reverse reaction Clostridium formicaceticum
1.5.1.20 FADH2 flavoprotein, reduced acceptor in forward reaction Clostridium formicaceticum
1.5.1.20 additional information no direct activity with pyridine nucleotides: NADH, NADPH Clostridium formicaceticum
1.5.7.1 FAD contains 1.7 FAD per 237000 Da enzyme Clostridium formicaceticum