The inhibitor protein IF 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 IF, but bovine IF 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 IF inhibits both the hydrolytic and synthetic activities of the human ATP synthase and that the activity of human IF is regulated by the phosphorylation of Ser-14 of mature IF. Here, we have made both human and bovine IF 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 submitochondrial particles. Over a wide range of conditions, including physiological conditions, both human and bovine IF are potent inhibitors of ATP hydrolysis, with no effect on ATP synthesis. Also, substitution of Ser-14 with phosphomimetic aspartic and glutamic acids had no effect on inhibitory properties, and Ser-14 is not conserved throughout mammals. Therefore, it is unlikely that the inhibitory activity of mammalian IF is regulated by phosphorylation of this residue.