Lysosomal H+-translocating ATPase has a similar subunit structure to chromaffin granule H+-ATPase complex.
Subunit structure of the lysosomal H+-ATPase was investigated using cold inactivation, immunological cross-reactivity with antibodies against individual subunits of the H+-ATPase from chromaffin granules and chemical modification with N,N'-dicyclohexyl[14C]carbodiimide. The lysosomal H+-ATPase was irreversibly inhibited when incubated at 0 degrees C in the presence of chloride or nitrate and MgATP. Inactivation in the cold resulted in the release of several polypeptides (72, 57, 41, 34 and 33 kDa) from the membrane, which had the same electrophoretic mobility as the corresponding subunits of chromaffin granule H+-ATPase. Cross-reactivity of antibodies revealed that the 72, 57 and 34 kDa polypeptides were immunologically identical to the corresponding subunits of chromaffin granule H+-ATPase. Dicyclohexylcarbodiimide, which inhibits proton translocation in the vacuolar ATPase, predominantly labeled two polypeptides of 18 and 15 kDa, which compose the membrane sector of the enzyme. These results suggest that the lysosomal H+-ATPase is a multimeric enzyme, whose subunit structure is similar to the chromaffin granule H+-ATPase. The subunit structure of other vacuolar H+-ATPases, revealed by cold inactivation and immunological cross-reactivity, is also presented.
Moriyama
Y
Y
Roche Institute of Molecular Biology, Roche Research Center, Nutley, NJ 07110.
Nelson
N
N
D000311:Adrenal Glands / Q000201:enzymology; D000345:Affinity Labels; D000818:Animals; D000906:Antibodies / Q000032:analysis; D002417:Cattle; D002837:Chromaffin Granules / Q000201:enzymology* / Q000276:immunology; D002838:Chromaffin System / Q000201:enzymology*; D003080:Cold Temperature; D003429:Cross Reactions; D004024:Dicyclohexylcarbodiimide / Q000032:analysis; D004591:Electrophoresis, Polyacrylamide Gel; D008099:Liver / Q000201:enzymology; D008247:Lysosomes / Q000201:enzymology* / Q000276:immunology; D010455:Peptides / Q000032:analysis; D006180:Proton-Translocating ATPases / Q000032:analysis*; D051381:Rats; D011919:Rats, Inbred Strains
D000345:Affinity Labels; D000906:Antibodies; D010455:Peptides; D004024:Dicyclohexylcarbodiimide; D006180:Proton-Translocating ATPases