Isolation and Characterization of DL-methylmalonyl-coenzyme A Racemase From Rat Liver

Arch Biochem Biophys. 1985 Aug 15;241(1):252-64. doi: 10.1016/0003-9861(85)90381-9.

Abstract

Certain amino acids and other compounds are metabolized via propionyl-CoA----D-methylmalonyl-CoA----L-methylmalonyl- CoA----succinyl-CoA----tricarboxylic acid cycle. D-Methylmalonyl-CoA can also be converted to methylmalonic acid and coenzyme A by a specific hydrolase that does not act on L-methylmalonyl-CoA [R.J. Kovachy, S.D. Copley, and R.H. Allen (1983) J. Biol. Chem. 258, 11415-11421]. Because little is known about mammalian DL-methylmalonyl-CoA racemase and because it is involved in the flow of D-methylmalonyl-CoA to L-methylmalonyl-CoA----tricarboxylic acid cycle (versus to methylmalonic acid), we developed a new assay and purified rat liver racemase 23,000-fold to homogeneity. The molecular weight of the racemase is 32,000 and it contains two subunits of Mr 16,000 that are not connected by disulfide bonds. The rat liver and the rat and human white blood cell racemase are immunologically related. They are completely inactivated by EDTA and can be activated by the addition of Co+2, with 50% activation occurring at a concentration of 0.2 microM. Lower levels for maximal activation were obtained with higher concentrations of Co+3, Fe+2, and Mn+2. Other metals such as Zn+2, Cu+2, Cu+1, and Cd+2 completely inhibited racemase even in the presence of equal concentrations of Co+2. The purified racemase appears to bind 1 mol Co/mol subunit.

Publication types

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

MeSH terms

  • Amino Acids / analysis
  • Animals
  • Cobalt / metabolism
  • Enzyme Activation / drug effects
  • Humans
  • Isomerases / isolation & purification*
  • Leukocytes / enzymology*
  • Liver / enzymology*
  • Metals / pharmacology
  • Methylmalonyl-CoA Mutase / isolation & purification*
  • Methylmalonyl-CoA Mutase / metabolism
  • Molecular Weight
  • Rats

Substances

  • Amino Acids
  • Metals
  • Cobalt
  • Isomerases
  • Methylmalonyl-CoA Mutase