ATP binding and hydrolysis steps of the uni-site catalysis by the mitochondrial F1-ATPase are affected by inorganic phosphate

The effect of inorganic phosphate (Pi) on uni-site ATP binding and hydrolysis by the nucleotide-depleted F1-ATPase from beef heart mitochondria (ndMF1) has been investigated. It is shown for the first time that Pi decreases the apparent rate constant of uni-site ATP binding by ndMF1 3-fold with the Kd of 0.38 ± 0.14 mM. During uni-site ATP hydrolysis, Pi also shifts equilibrium between bound ATP and ADP + Pi in the direction of ATP synthesis with the Kd of 0.17 ± 0.03 mM. However, 10 mM Pi does not significantly affect ATP binding during multi-site catalysis.

Research Highlights►Phosphate decreases the rate of ATP binding by mitochondrial F1-ATPase under uni-site conditions. ►Phosphate shifts an equilibrium at the catalytic site during uni-site ATP hydrolysis in the direction of ATP synthesis. ►Phosphate does not affect multi-site ATP hydrolysis.