Clathrin-coated vesicles from rat liver: Enzymatic profile and characterization of ATP-dependent proton transport

Clathrin-coated vesicles isolated from rat liver exhibited an enzymatic profile distinct from that of rat liver plasma membranes, lysosomes, microsomes, and mitochondria. The coated vesicles catalyzed ATP-dependent proton transport that acidified the vesicle interior, as measured by the fluorescence quenching of acridine orange. H⁺ transport by coated vesicles was not inhibited by vanadate (0.1 mM) or ouabain (2 mM) and differed from H⁺ transport by rat liver submitochondrial particles in its greater resistance to inhibition by oligomycin (10 pM to 10 μM) and N,N'-dicyclohexylcarbodiimide (DCCD) (0.1-100 μM) and its sensitivity to N-ethylmaleimide (0.1-2 mM). H⁺ transport was stimulated by valinomycin in the presence of K⁺, exhibited no specific cation requirement, but was dependent upon the presence of a permeant anion, with Cl^- and Br^- being the most effective of the anions studied. Finally, H⁺ transport was poorly supported by GTP, UTP, or ADP and exhibited no consistent relationship to the coated vesicle-associated ouabain-insensitive ATPase activity.