PMID:12595534
| Citation |
Weber, J, Muharemagic, A, Wilke-Mounts, S and Senior, AE (2003) F1F0-ATP synthase. Binding of delta subunit to a 22-residue peptide mimicking the N-terminal region of alpha subunit. J. Biol. Chem. 278:13623-6 |
|---|---|
| Abstract |
The stator in F(1)F(0)-ATP synthase resists strain generated by rotor torque. In Escherichia coli the b(2)delta subunit complex comprises the stator, bound to subunit a in F(0) and to alpha(3)beta(3) hexagon of F(1). Proteolysis and cross-linking had suggested that N-terminal residues of alpha subunit are involved in binding delta. Here we demonstrate that a synthetic peptide consisting of the first 22 residues of alpha ("alpha N1-22") binds specifically to isolated wild-type delta subunit with high affinity (K(d) = 130 nm), accounting for a major portion of the binding energy when delta-depleted F(1) and isolated delta bind together (K(d) = 1.4 nm). Stoichiometry of binding of alpha N1-22 to delta at saturation was 1/1, showing that in intact F(1)F(0)-ATP synthase only one of the three alpha subunits is involved in delta binding. When alpha N1-22 was incubated with delta subunits containing mutations in helices 1 or 5 on the F(1)-binding face of delta, peptide binding was impaired as was binding of delta-depleted F(1). Residues alpha 6-18 are predicted to be helical, and a potential helix capping box occurs at residues alpha 3-8. Circular dichroism measurements showed that alpha N1-22 had significant helical content. Hypothetically a helical region of residues alpha N1-22 packs with helices 1 and 5 on the F(1)-binding face of delta, forming the alpha/delta interface. |
| Links |
PubMed Online version:10.1074/jbc.C300061200 |
| Keywords |
Circular Dichroism; Dose-Response Relationship, Drug; Escherichia coli/enzymology; Hydrogen-Ion Concentration; Ions; Kinetics; Magnesium/pharmacology; Mutation; Peptides/chemistry; Protein Binding; Protein Structure, Tertiary; Proton-Translocating ATPases/chemistry; Proton-Translocating ATPases/metabolism; Spectrometry, Fluorescence |
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