Mobilization of iron from endocytic vesicles. The effects of acidification and reduction

Abstract

The factors necessary to dissociate iron from transferrin in endocytic vesicles and to mobilize the iron across the vesicle membrane were studied in a preparation of endocytic vesicles markedly enriched in transferrin-transferrin receptor complexes isolated from rabbit reticulocytes. Vesicles were prepared with essentially fully saturated transferrin by incubating the reticulocytes with the protonophore carbonyl cyanide 4-(trifluoromethoxy)phenylhydrazone prior to incubation with 59Fe,125I-transferrin with or without fluorescein isothiocyanate labeling. Initiation of acidification by the addition of ATP was sufficient to achieve dissociation of 59Fe from transferrin with a rate constant of 0.054 ± 0.06 s-1. Mobilization of 59Fe out of the vesicles required, besides ATP, the addition of a reductant with 1 mM ascorbate, allowing ~60% mobilization at 10 min with a rate constant of 0.0038 ± 0.0006 s-1. An NADH:ferricyanide reductase activity could be demonstrated in the vesicles with an ac