title: Interaction of adenine nucleotides with multiple binding sites on beef heart mitochondrial adenosine triphosphatase.
abstract: Beef heart mitochondrial ATPase (F1) contained 2 mol of ADP and 1 mol of ATP/mol of enzyme, which resisted removal by Sephadex chromatography with dilute buffers or repeated precipitation with ammonium sulfate. The native enzyme also contained two apparently equivalent binding sites, which participated in readily reversible binding of adenyl-5'-ylimidodiphosphate (AMP-P(NH)P), with a Kd of 1.3 mum. The failure of AMP-P(NH)P to compete effectively with ADP for binding sites on F1 may be related to the failure of the analog to inhibit oxidative phosphorylation. Virtually complete removal of all adenine nucleotides from F1 occurred when the enzyme was chromatographed on columns of Sephadex equilibrated with 50% glycerol. No loss in ATPase activity was observed following removal of nucleotides from the enzyme, which was then capable of binding more than 4 mol of ADP and almost 5 mol of AMP-P(NH)P/mol of protein. Subsequent chromatography on columns of Sephadex equilibrated with dilute buffers containing Mg2+ removed only 1.5 mol of ADP and no AMP-P(NH)P from the enzyme. Reconstitution of F1 with ADP or with almost 5 mol of AMP-P(NH)P resulted in preparations that exhibited an undiminished capacity to restore oxidative phosphorylation in F1-deficient submitochondrial particles.
mesh_terms: D000227:Adenine Nucleotides; D000244:Adenosine Diphosphate; D000251:Adenosine Triphosphatases / Q000378:metabolism; D000255:Adenosine Triphosphate; D000266:Adenylyl Imidodiphosphate; D000818:Animals; D001665:Binding Sites; D002417:Cattle; D004355:Drug Stability; D005990:Glycerol; D007700:Kinetics; D008931:Mitochondria, Muscle / Q000201:enzymology*; D009206:Myocardium; D010085:Oxidative Phosphorylation; D011485:Protein Binding; D011489:Protein Denaturation
chemical_list: D000227:Adenine Nucleotides; D000266:Adenylyl Imidodiphosphate; D000244:Adenosine Diphosphate; D000255:Adenosine Triphosphate; D000251:Adenosine Triphosphatases; D005990:Glycerol