The inhibitor protein IF1 from mammalian mitochondria inhibits ATP hydrolysis but not ATP synthesis by the ATP synthase complex

The hydrolytic activity of the ATP synthase in bovine mitochondria is inhibited by a protein called IF1, but bovine IF1 has no effect on the synthetic activity of the bovine enzyme in mitochondrial vesicles in the presence of a proton motive force. In contrast, it has been suggested based on indirect observations that human IFI inhibits both the hydrolytic and synthetic activities of the human ATP synthase, and that the activity of human IF1 is regulated by the phosphorylation of serine-14 of mature IF1. Here, we have made both human and bovine IF1 which are 81 and 84 amino acids long, respectively, and identical in 71.4% of their amino acids, and have investigated their inhibitory effects on the hydrolytic and synthetic activities of ATP synthase in bovine sub-mitochondrial particles. Over a wide range of conditions, including physiological conditions, both human and bovine IF1 are potent inhibitors of ATP hydrolysis, with no effect on ATP synthesis. Also, substitution of serine-14 with phosphomimetic aspartic and glutamic acids had no effect on inhibitory properties, and serine-14 is not conserved throughout mammals. Therefore, it is unlikely that the inhibitory activity of mammalian IF1 is regulated by phosphorylation of this residue.