Nucleotide-CF1 interactions and current views on the catalytic mechanism

The binding of nucleotides on isolated subunits as well as on reconstituted CF1 core complex is reviewed. Nucleotide interaction with CF1 and conseque nt ATPase activity are always associated with the presence of Mg2+. The met al binding site studies using Electron Paramagnetic Resonance (EPR) and pul sed EPR conclude that the metal binding occurs prior to any nucleotide addi tion. The addition of nucleotide does not modify the enzyme's metal binding site but brings on additional ligands with the phosphates of the nucleotid es. The ATPase and nucleotide binding experiments with CF1 are also better interpreted by the hypothesis that Mg2+ is an activator rather than an inhi bitor of the enzyme and that the actual substrate of CF1-ATPase is ATP rath er than MgATP. The dual role of tentoxin as an inhibitor at low concentrati on (10(-8)-10(-7) M) and activator at higher concentrations (10(-6) M) of t he enzymatic activity of CF1, is due to the presence of two different bindi ng sites on CF1. The synthesis of a new cyclic analogue of tentoxin with al anine changed for a serine has shown that it was possible to dissociate the two roles. The serine tentoxin analogue has the same inhibition effect on CF1 but is no longer an activator. The binding of nucleotides may influence the stability, produce structural changes and, over long distance, cause m ovements of CF1. All these effects of nucleotide or metal binding and activ ation or inhibition of CF1 may help also to elucidate the role played by th e catalytic and non catalytic sites. These questions are reviewed and analy zed with respect to the current views on the catalytic mechanism.