(1) Incubation of the beef heart mitochondrial ATPase, F1with Mg-ATP was required for the binding of the natural inhibitor, IF1, to F1to form the inactive F1-IF1complex. When F1was incubated in the presence of [14C]ATP and MgCl2, about 2 mol14C-labeled adenine nucleotides were found to bind per mol of F1; the bound14C-labeled nucleotides consisted of [14C]ADP arising from [14C]ATP hydrolysis and [14C]ATP. The14C-labeled nucleotide binding was not prevented by IF1. These data are in agreement with the idea that the formation of the F1-IF1complex requires an appropriate conformation of F1. (2) The14C-labeled adenine nucleotides bound to F1following preincubation of F1with Mg-[14C]ATP could be exchanged with added [3H]ADP or [3H]ATP. No exchange occurred between added [3H]ADP or [3H]ATP and the14C-labeled adenine nucleotides bound to the F1-IF1complex. These data suggest that the conformation of F1in the isolated F1-IF1complex is further modified in such a way that the bound14C-labeled nucleotides are no longer available for exchange. (3)32Piwas able to bind to isolated F1with a stoichiometry of about 1 mol of Piper mol of F1(Penefsky, H.S. (1977) J. Biol. Chem. 252, 2891-2899). There was no binding of32Pito the F1-IF1complex. Thus, not only the nucleotides sites, but also the Pisite, are masked from interaction with external ligands in the isolated F1-IF1complex. © 1981.
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