A case study in applying docking predictions: Modelling the tentoxin binding sites of chloroplast F1-ATPase

The study presents an ab initio approach for locating a ligand-binding site and demonstrates that relevant conclusions can be deduced from multiple, predicted ligand positions. Tentoxin is a specific inhibitor of plastid CF1-ATPase, its interaction with the α- and β-subunits of the enzyme resulting in complex interference with ATPase activity. The uniquely different conformations of the β-subunits in the quasisymmetrical structure of the F1-ATPase from bovine mitochondria offer an opportunity to model tentoxin binding at multiple sites in plastid CF1-ATPase. Using software for molecular docking, we located and analysed three putative binding sites with approximately equally high complementarity to tentoxin. Complementarity at these sites is sensitive to the nucleotide occupancy state of the β-subunit. The main interactions stabilizing the putative complexes were determined, and homology models for the tentoxin-binding sites of Chlamydomonas plastid CF1-ATPase were created. The predicted binding pocket residues for Site I are at the α_TP/β_TP interface and include residue Glu-67β_TP (codon 83β in plastid CF1-ATPase), previously identified as a molecular-genetic determinant for the high affinity, inhibitory response to tentoxin in Chlamydomonas. This site overlaps with that suggested by others, but ∼ 50% of residues differ. Sites II and III, possibly related to low-affinity binding of tentoxin, are located in functionally active regions: one entirely within the α_TP-subunit and sharing a residue with the α_TP nucleotide-binding site; the other at the α_TP/β_E/γ interface and sharing a residue with the conserved DELSEED sequence. Non-catalytic residues in these putative pockets represent potential targets for mutational analysis.