Cobalamin Binding and Cobalamin-Dependent Enzyme Activity in Normal and Mutant Human Fibroblasts
AUTOR(ES)
Mellman, Ira
RESUMO
We have studied the intracellular binding of radioactive cobalamin by normal cultured human fibroblasts grown in medium containing [57Co]-cobalamin. We have also assessed the significance of defects in this binding activity exhibited by two classes of human mutants (cbl C and cbl D) each characterized by pleiotropic deficiencies in the accumulation and retention of cobalamin, in the synthesis of cobalamin coenzymes, and accordingly, in the holoenzyme activities of both cobalamin-dependent enzymes, 5-methyltetrahydrofolate:homocysteine methyltransferase and methylmalonyl-CoA mutase. Based on the coincidence of [57Co]cobalamin binding and cobalamin-dependent enzyme activities after Sephadex G-150 chromatography and polyacrylamide gel electrophoresis, we conclude that, as in rat liver, the intracellular binding of labeled cobalamin by normal fibroblasts reflects the attachment of the vitamin to the cobalamin-dependent methyltransferase and mutase. Whereas cbl C cells are completely deficient in the binding of [57Co]cobalamin to either enzyme, fibroblasts which bear the phenotypically similar but genetically distinct cbl D mutation retain some binding activity, and accordingly, have higher holomethyltransferase and holomutase activities than do cbl C cells. The defect in [57Co]-cobalamin binding exhibited by both cbl C and cbl D fibroblasts is almost certainly not a result of mutations which affect the methyltransferase or mutase apoenzymes, since the electrophoretic mobilities and the affinities of these enzymes for their respective cobalamin coenzymes are indistinguishable from those in control cell extracts. These results suggest that both the cbl C and cbl D mutations affect some enzymatic step(s) which converts newly taken up cobalamin to a form capable of being bound by the two cobalamin-dependent enzymes.
ACESSO AO ARTIGO
http://www.pubmedcentral.nih.gov/articlerender.fcgi?artid=371853Documentos Relacionados
- Cobalamins and cobalamin-dependent enzymes in Candida utilis.
- Inhibition of cobalamin-dependent enzymes by cobalamin analogues in rats.
- Intracellular binding of radioactive hydroxocobalamin to cobalamin-dependent apoenzymes in rat liver.
- Coupled Changes in Translation and Transcription during Cobalamin-Dependent Regulation of btuB Expression in Escherichia coli
- Mapping the interactions between flavodoxin and its physiological partners flavodoxin reductase and cobalamin-dependent methionine synthase