Van Dyke R W, Steer C J, Scharschmidt B F
Proc Natl Acad Sci U S A. 1984 May;81(10):3108-12. doi: 10.1073/pnas.81.10.3108.
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 microM) and N,N'-dicyclohexylcarbodiimide (DCCD) (0.1-100 microM) 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.
从大鼠肝脏分离出的网格蛋白包被小泡呈现出与大鼠肝脏质膜、溶酶体、微粒体和线粒体不同的酶学特征。通过吖啶橙荧光猝灭测量发现，包被小泡催化依赖ATP的质子转运，使小泡内部酸化。包被小泡的H⁺转运不受钒酸盐（0.1 mM）或哇巴因（2 mM）的抑制，并且与大鼠肝脏亚线粒体颗粒的H⁺转运不同，它对寡霉素（10 pM至10 μM）和N,N'-二环己基碳二亚胺（DCCD）（0.1 - 100 μM）的抑制具有更大的抗性，对N-乙基马来酰亚胺（0.1 - 2 mM）敏感。在有K⁺存在的情况下，缬氨霉素刺激H⁺转运，对特定阳离子无需求，但依赖于渗透性阴离子的存在，在所研究的阴离子中，Cl⁻和Br⁻是最有效的。最后，GTP、UTP或ADP对H⁺转运的支持作用较弱，并且与包被小泡相关的哇巴因不敏感ATP酶活性没有一致的关系。
