The proton adenosinetriphosphatase complex of rat liver mitochondria. Temperature-dependent dissociation-reassociation of the F1-ATPase subunits.
The soluble F1 moiety of the rat liver mitochondrial proton ATPase dissociates into two easily separable fractions when cold treated and then warmed. One fraction is soluble in potassium phosphate buffer, pH 7.4, whereas the other is insoluble. Neither of these two fractions alone can catalyze ATP hydrolysis under assay conditions optimal for the native F1-ATPase. The insoluble fraction when resolved via sodium dodecyl sulfate--polyacrylamide gel electrophoresis is shown to be composed of only alpha and gamma subunits. When this fraction is chromatographed on Sephadex G-75, it is resolved into an alpha gamma complex and into the alpha subunit alone. The soluble fraction when resolved in the same electrophoretic system is shown to contain the remaining subunits, beta, delta, epsilon, and some gamma. This fraction is resolved into two major components by chromatography on Sepharose CL-6B, a beta gamma complex and beta subunit alone. The cold-dissociated enzyme can be readily associated when the temperature is raised to 20 degrees C. In the presence of either ATP or MgATP the enzyme completely regains its original ATPase specific activity. In contrast, Mg2+ is only about 15% effective in restoring ATPase activity. The results presented here define conditions for the dissociation and reassociation of the major subunits comprising the F1-ATPase of rat liver and thus provide a unique system among mammalian enzymes for testing the function of individual subunits. In addition, they strongly indicate that neither the alpha nor beta subunits, nor complexes of these subunits with the gamma subunit, are capable of catalyzing ATP hydrolysis.(ABSTRACT TRUNCATED AT 250 WORDS)
D000255:Adenosine Triphosphate / Q000494:pharmacology; D000818:Animals; D066298:In Vitro Techniques; D008274:Magnesium / Q000494:pharmacology; D008297:Male; D008930:Mitochondria, Liver / Q000201:enzymology*; D011487:Protein Conformation / Q000187:drug effects; D006180:Proton-Translocating ATPases / Q000378:metabolism*; D051381:Rats; D013696:Temperature
D000255:Adenosine Triphosphate; D006180:Proton-Translocating ATPases; D008274:Magnesium