Properties of a novel ATPase enzyme in chromaffin granules.
Membranes were isolated from mitochondria and chromaffin granules of bovine adrenal medullae. The cross-contamination between the two membranes was examined by comparing the radioactive bands on autoradiograms of gels after phosphorylation of the membranes with [gamma-32P]-ATP and decoration with [125I]concanavalin A and [125I]protein A with antibody that was raised against chromaffin-granule membranes. It was found that the membranes cross-contaminated each other by less than 10%. The technique of immunodecoration with antibodies against beta subunits of proton-ATPases from yeast mitochondria, spinach chloroplasts, and E. coli membranes was used for quantitative estimation of proton-ATPase complexes in chromaffin granules and mitochondrial membranes. It was found that chromaffin-granule membranes contain less than 10% of the amount of proton-ATPase complex in mitochondrial membranes. The specific ATPase activity of chromaffin-granule membranes was on the order of 30 to 50% of the mitochondrial membranes. The ATPase activity of the chromaffin-granule membranes was more sensitive to 4-acetamido-4'-isothiocyano-2,2'-disulfonic acid stilbene and 4-chloro-7-nitrobenzofurazan. It was much less sensitive than the mitochondrial membranes to antibody against beta subunit of proton-ATPase from E. coli membranes. After solubilization of chromaffin-granule membranes by octyglucoside and cholate and subsequent centrifugation on sucrose gradient, two different ATPase enzymes were separated. The heavier enzyme was identical to the mitochondrial-ATPase complex, while the lighter enzyme was identified as a novel ATPase, which might be responsible for the special properties of the ATPase activity of chromaffin-granule membranes.
D000251:Adenosine Triphosphatases / Q000378:metabolism*; D000313:Adrenal Medulla / Q000201:enzymology; D000818:Animals; D001345:Autoradiography; D002417:Cattle; D002837:Chromaffin Granules / Q000201:enzymology*; D002838:Chromaffin System / Q000201:enzymology*; D003208:Concanavalin A / Q000494:pharmacology; D004926:Escherichia coli / Q000201:enzymology; D007120:Immunochemistry; D066298:In Vitro Techniques; D010085:Oxidative Phosphorylation
D003208:Concanavalin A; D000251:Adenosine Triphosphatases