Similarity of lysosomal H⁺-ATPase to mitochondrial F₀F₁-ATPase in sensitivity to anions and drugs as revealed by solubilization and reconstitution

Lysosomal H⁺-translocating ATPase (H⁺-ATPase) was solubilized with lysophosphatidylcholine and reconstituted into liposomes (Moriyama, Y., Takano, T. and Ohkuma, S. (1984) J. Biochem. (Tokyo) 96, 927-930). In this study, the sensitivities of membrane-bound, solubilized and liposome-incorporated ATPase to various anions and drugs were measured in comparison with those of similar forms of mitochondrial H⁺-ATPase (mitochondrial F₀F₁-ATPase) with the following results. (1) Bicarbonate and sulfite activated solubilized lysosomal H⁺-ATPase, but not the membrane-bound ATPase or ATPase incorporated into liposomes. All three forms of mitochondrial F₀F₁-ATPase were activated by these anions. (2) All three forms of both lysosomal H⁺-ATPase and mitochondrial F₀F₁-ATPase were strongly inhibited by SCN^-, NO₃^- and F^-, but scarcely affected by Cl^-, Br^- and SO₄^2-. (3) The solubilized lysosomal H⁺-ATPase was strongly inhibited by azide, quercetin, 7-chloro-4-nitrobenzo-2-oxa-1,3-diazole (NBD-Cl), 4-acetamido-4′-isothiocyanatostilbene-2,2′-disulfonic acid (SITS), 4,4′-diisothiocyanatostilbene-2,2′-disulfonic acid (DIDS) and oligomycin. Its sensitivity was almost the same as that of mitochondrial F₀F₁-ATPase. Neither membrane-bound ATPase nor ATPase incorporated into liposomes was affected appreciably by these drugs. These results indicate that the sensitivity to anions and drugs of lysosomal H⁺-ATPase depends on the form of the enzyme and that the sensitivity of the solubilized lysosomal H⁺-ATPase is very similar to that of mitochondrial F₀F₁-ATPase. On the other hand, the two ATPases differ in their sensitivity to N-ethylmaleimide and pyridoxal phosphate: only the mitochondrial ATPase is inhibited by pyridoxal phosphate whereas only the lysosomal ATPase is inhibited by N-ethylmaleimide.