IDENTIFICATION AND CHARACTERIZATION OF 2 ENZYMES INVOLVED IN THE INTRACELLULAR METABOLISM OF COBALAMIN - CYANOCOBALAMIN BETA-LIGAND TRANSFERASE AND MICROSOMAL COB(III) ALAMIN REDUCTASE
Eh. Pezacka, IDENTIFICATION AND CHARACTERIZATION OF 2 ENZYMES INVOLVED IN THE INTRACELLULAR METABOLISM OF COBALAMIN - CYANOCOBALAMIN BETA-LIGAND TRANSFERASE AND MICROSOMAL COB(III) ALAMIN REDUCTASE, Biochimica et biophysica acta, 1157(2), 1993, pp. 167-177
Two enzymes involved in the intracellular metabolism of cobalamin have
been identified and characterized: cyanocobalamin beta-ligand transfe
rase and microsomal cob(III)alamin reductase. The beta-ligand transfer
ase is a cytosolic enzyme utilizing FAD, NADPH and reduced glutathione
. The product of the reaction has been identified as glutathionylcobal
amin. NADH-linked cob(III)alamin reductase has been found in two subce
llular fractions: microsomal and inner mitochondrial membrane. The pro
duct of the reduction catalyzed by the microsomal enzyme has been iden
tified as cob(II)alamin. In cbl C mutant fibroblasts, the specific act
ivities of cyanocobalamin beta-ligand transferase and cob(III)alamin r
eductase were markedly decreased and have varied from 3%-30% and 36%-4
2% of normal, respectively. The specific activity of mitochondrial cob
(III)alamin reductase was only 30% of normal in two cbl C mutants and
normal in remaining mutant cell lines. In the cbl D cells, the specifi
c activities were 33% and 55%. Mitochondrial cob(III)alamin reductase
was not affected by cbl D mutation. Methionine synthase, L-methylmalon
yl-CoA mutase and microsomal cytochrome c and b5 reductases are not af
fected by both mutations. The cbl E mutation affects only the activity
of methionine synthase. These results support the hypothesis that the
early enzymatic steps of intracellular metabolism of cobalamin are si
milar in the synthesis of both methylcobalamin and adenosylcobalamin a
nd these steps are altered by the cbl C and cbl D mutations.