Purification and properties of 5,10-methylenetetrahydrofolate reductase, an iron-sulfur flavoprotein from Clostridium formicoaceticum

J Biol Chem. 1984 Sep 10;259(17):10845-9.


Methylenetetrahydrofolate reductase in Clostridium formicoaceticum has been purified to a specific activity of 140 mumol min-1 mg-1 when assayed at 37 degrees C, pH 7.2, in the direction of oxidation of 5-methyltetrahydrofolate with benzyl viologen as electron acceptor. The purified enzyme is judged to be homogeneous by polyacrylamide disc-gel electrophoresis and gel filtration. The enzyme which is an octamer has a molecular weight of about 237,000 and consists of four each of two different subunits having the molecular weights 26,000 and 35,000. The octameric enzyme contains per mol 15.2 +/- 0.3 iron, 2.3 +/- 0.2 zinc, 19.5 +/- 1.3 acid-labile sulfur, and 1.7 FAD. The UV-visible absorbance spectrum has a peak at 385 nm and a shoulder at 430 nm and is that of a flavoprotein containing iron-sulfur centers. The reductase, which is sensitive to oxygen, must be handled anaerobically and is stabilized by 2 mM dithionite. It catalyzes the reduction of methylene blue, menadione, benzyl viologen, rubredoxin, and FAD with 5-methyltetrahydrofolate and the oxidation of reduced ferredoxin and FADH2 with 5,10-methylenetetrahydrofolate. No activity was observed with pyridine nucleotides. It is suggested that the physiologically important reaction catalyzed by the enzyme is the reduced ferredoxin-dependent reduction of 5,10-methylenetetrahydrofolate to 5-methyltetrahydrofolate.

Publication types

  • Research Support, U.S. Gov't, P.H.S.

MeSH terms

  • 5,10-Methylenetetrahydrofolate Reductase (FADH2)
  • Alcohol Oxidoreductases / isolation & purification*
  • Alcohol Oxidoreductases / metabolism
  • Clostridium / enzymology*
  • Iron-Sulfur Proteins / isolation & purification
  • Kinetics
  • Macromolecular Substances
  • Molecular Weight
  • Substrate Specificity


  • Iron-Sulfur Proteins
  • Macromolecular Substances
  • Alcohol Oxidoreductases
  • 5,10-Methylenetetrahydrofolate Reductase (FADH2)