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Literature summary for 1.5.1.20 extracted from
- Purification and properties of 5,10-methylenetetrahydrofolate reductase, an iron-sulfur flavoprotein from Clostridium formicoaceticum (1984), J. Biol. Chem., 259, 10845-10849.
General Stability
| General Stability | Organism |
|---|---|
| 0.005 mM FAD stabilizes enzyme during purification procedure | Clostridium formicaceticum |
KM Value [mM]
| KM Value [mM] | KM Value Maximum [mM] | Substrate | Comment | Organism | Structure |
|---|---|---|---|---|---|
| 0.12 | - |
5-methyltetrahydrofolate | - |
Clostridium formicaceticum |  |
| 11.1 | - |
benzyl viologen | - |
Clostridium formicaceticum | |
Metals/Ions
| Metals/Ions | Comment | Organism | Structure |
|---|---|---|---|
| Iron | 15.2 molecules iron per enzyme molecule, enzyme contains iron-sulfur clusters | Clostridium formicaceticum | |
| iron-sulfur centre | enzyme contains 15.2 molecules iron and 19.5 molecules acid-labile sulfur as iron-sulfur clusters | Clostridium formicaceticum | |
| Zinc | 2.3 molecules zinc per enzyme molecule | Clostridium formicaceticum | |
Molecular Weight [Da]
| Molecular Weight [Da] | Molecular Weight Maximum [Da] | Comment | Organism |
|---|---|---|---|
| 26000 | - |
alpha4,beta4, 4 * 26000 + 4 * 35000, SDS-PAGE | Clostridium formicaceticum |
| 26000 | - |
alpha2,beta2, 2 * 26000 + 2 * 35000, determined at pH 9, SDS-PAGE | Clostridium formicaceticum |
| 35000 | - |
alpha4,beta4, 4 * 26000 + 4 * 35000, SDS-PAGE | Clostridium formicaceticum |
| 35000 | - |
alpha2,beta2, 2 * 26000 + 2 * 35000, determined at pH 9, SDS-PAGE | Clostridium formicaceticum |
| 124000 | - |
determination at pH 9, alpha2,beta2 enzyme form, PAGE | Clostridium formicaceticum |
| 237000 | - |
gel filtration | Clostridium formicaceticum |
| 237000 | - |
determination at pH 7.4, alpha4,beta4 enzyme form | Clostridium formicaceticum |
Natural Substrates/ Products (Substrates)
| Natural Substrates | Organism | Comment (Nat. Sub.) | Natural Products | Comment (Nat. Pro.) | Rev. | Reac. |
|---|---|---|---|---|---|---|
| 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 | - |
? | |
| 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 | - |
? | |
| 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
| Organism | UniProt | Comment | Textmining |
|---|---|---|---|
| Clostridium formicaceticum | - |
strain ATCC 23439 | - |
Oxidation Stability
| Oxidation Stability | Organism |
|---|---|
| oxygen labile, half-life: less than 1 h in aerobic buffer, sodium dithionite prevents inactivation by oxygen | Clostridium formicaceticum |
Purification (Commentary)
| Purification (Comment) | Organism |
|---|---|
- |
Clostridium formicaceticum |
Specific Activity [micromol/min/mg]
| Specific Activity Minimum [µmol/min/mg] | Specific Activity Maximum [µmol/min/mg] | Comment | Organism |
|---|---|---|---|
| 139 | - |
- |
Clostridium formicaceticum |
Storage Stability
| Storage Stability | Organism |
|---|---|
| 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 |
| frozen, 50 mM Tris-HCl buffer, pH 7.4, 20% glycerol, 2 mM dithionite, longer time, 10-20% loss of activity | Clostridium formicaceticum |
Substrates and Products (Substrate)
| Substrates | Comment Substrates | Organism | Products | Comment (Products) | Rev. | Reac. |
|---|---|---|---|---|---|---|
| 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 | - |
? | |
| 5,10-methylenetetrahydrofolate + reduced acceptor | forward reaction: reduced acceptor is FADH2 | Clostridium formicaceticum | 5-methyltetrahydrofolate + oxidized acceptor | - |
r | |
| 5,10-methylenetetrahydrofolate + reduced acceptor | reverse reaction: menadione as electron acceptor | Clostridium formicaceticum | 5-methyltetrahydrofolate + oxidized acceptor | - |
r | |
| 5,10-methylenetetrahydrofolate + reduced acceptor | reverse reaction: FAD, rubredoxin, benzyl viologen and methylene blue as electron acceptor | Clostridium formicaceticum | 5-methyltetrahydrofolate + oxidized acceptor | - |
r | |
| 5,10-methylenetetrahydrofolate + reduced acceptor | forward reaction: reduced ferredoxin as reduced acceptor | Clostridium formicaceticum | 5-methyltetrahydrofolate + oxidized acceptor | - |
r | |
| 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 | - |
? | |
| 5,10-methylenetetrahydrofolate + reduced ferredoxin | physiological important reaction: enzyme catalyzes reduction of 5,10-methylenetetrahydrofolate with reduced ferredoxin | Clostridium formicaceticum | 5-methyltetrahydrofolate + oxidized ferredoxin | - |
? | |
| 5-methyltetrahydrofolate + NAD+ | - |
Clostridium formicaceticum | 5,10-methylenetetrahydrofolate + NADH + H+ | - |
r | |
| additional information | no direct activity with pyridine nucleotides | Clostridium formicaceticum | ? | - |
? | |
| nitrate + reduced benzyl viologen | - |
Clostridium formicaceticum | nitrite + benzyl viologen | - |
? | |
Subunits
| Subunits | Comment | Organism |
|---|---|---|
| octamer | alpha4,beta4, 4 * 26000 + 4 * 35000, SDS-PAGE | Clostridium formicaceticum |
| octamer | determined at pH 7.4 | Clostridium formicaceticum |
| tetramer | alpha2,beta2, 2 * 26000 + 2 * 35000, determined at pH 9, SDS-PAGE | Clostridium formicaceticum |
Temperature Optimum [°C]
| Temperature Optimum [°C] | Temperature Optimum Maximum [°C] | Comment | Organism |
|---|---|---|---|
| 37 | - |
assay at | Clostridium formicaceticum |
pH Optimum
| pH Optimum Minimum | pH Optimum Maximum | Comment | Organism |
|---|---|---|---|
| 7.8 | 8 | - |
Clostridium formicaceticum |
pH Stability
| pH Stability | pH Stability Maximum | Comment | Organism |
|---|---|---|---|
| 7.1 | 7.6 | at this pH-range enzyme is more stable in Tris-HCl than in triethanolamine-HCl or phosphate buffers | Clostridium formicaceticum |
Cofactor
| Cofactor | Comment | Organism | Structure |
|---|---|---|---|
| FAD | 1.7 molecules enzyme-bound FAD per enzyme molecule | Clostridium formicaceticum | |
| FAD | flavoprotein, oxidized acceptor in reverse reaction | Clostridium formicaceticum | |
| FADH2 | flavoprotein, reduced acceptor in forward reaction | Clostridium formicaceticum | |
| additional information | no direct activity with pyridine nucleotides: NADH, NADPH | Clostridium formicaceticum |
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