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 SO2-4. (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.
Moriyama
Y
Y
Takano
T
T
Ohkuma
S
S
D000818:Animals; D000838:Anions; D001639:Bicarbonates / Q000494:pharmacology; D002462:Cell Membrane / Q000201:enzymology; D004024:Dicyclohexylcarbodiimide / Q000494:pharmacology; D004789:Enzyme Activation / Q000187:drug effects; D005033:Ethylmaleimide / Q000494:pharmacology; D007700:Kinetics; D008099:Liver / Q000201:enzymology*; D008247:Lysosomes / Q000201:enzymology*; D008930:Mitochondria, Liver / Q000201:enzymology*; D006180:Proton-Translocating ATPases / Q000037:antagonists & inhibitors / Q000378:metabolism*; D011732:Pyridoxal Phosphate / Q000494:pharmacology; D051381:Rats; D012995:Solubility; D013447:Sulfites / Q000494:pharmacology
D000838:Anions; D001639:Bicarbonates; D013447:Sulfites; D004024:Dicyclohexylcarbodiimide; D011732:Pyridoxal Phosphate; D006180:Proton-Translocating ATPases; D005033:Ethylmaleimide