Uptake and metabolism of free cyanocobalamin by cultured human fibroblasts from controls and a patient with transcobalamin II deficiency.
We have investigated the uptake and metabolism of free cyanocobalamin (CN-Cbl; vitamin B12) by intact cultured human skin fibroblasts. Monolayers of control fibroblasts take up free CN-[57Co]Cbl via a saturable, calcium-independent process that is inhibited by sulfhydryl reagents, inhibitors of protein synthesis, and inhibitors of electron transport, but not by inhibitors of glycolysis. CN-Cbl taken up in this manner is converted to active cobalamin (Cbl) coenzymes (adenosylcobalamin and methylcobalamin) and becomes associated with intracellular Cbl-dependent apoenzymes (methylmalonyl CoA mutase and homocysteine:methyltetrahydrofolate methyltransferase). Since fibroblasts from controls were also found to synthesize transcobalamin II (TC II), a plasma protein shown previously to facilitate the cellular uptake of Cbl, it seemed possible that the observed uptake of free CN-Cbl was TC II-mediated. This thesis was rejected by demonstrating that cells from a patient with complete TC II deficiency took up free CN-Cbl as well as control cells did. Finally, we propose a mechanism by which an uptake process for free Cbl might serve a function in intracellular metabolism of Cbl.
D001798:Blood Proteins / Q000172:deficiency*; D002478:Cells, Cultured; D003513:Cycloheximide / Q000494:pharmacology; D005033:Ethylmaleimide / Q000494:pharmacology; D005347:Fibroblasts / Q000378:metabolism*; D006801:Humans; D007700:Kinetics; D011190:Potassium Cyanide / Q000494:pharmacology; D014155:Transcobalamins / Q000172:deficiency*; D014805:Vitamin B 12 / Q000378:metabolism*
D001798:Blood Proteins; D014155:Transcobalamins; D003513:Cycloheximide; D011190:Potassium Cyanide; D005033:Ethylmaleimide; D014805:Vitamin B 12