Analysis of sequence determinants of F1Fo-ATP synthase in the N-terminal region of α subunit for binding of δ subunit

Joachim Weber, Alma Muharemagic, Susan Wilke-Mounts, Alan E. Senior

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The stator in F1Fo-ATP synthase resists strain generated by rotor torque. In Escherichia coli, the b2δ subunit complex comprises the stator, bound to subunit α in Fo and to the α3β3 hexagon of F1. Previous work has shown that N-terminal residues of α subunit are involved in binding δ. A synthetic peptide consisting of the first 22 residues of α (αN1-22) binds specifically to isolated wild-type δ subunit with 1:1 stoichiometry and high affinity, accounting for a major portion of the binding energy between δ and F1. Residues α6-18 are predicted by secondary structure algorithms and helical wheels to be α-helical and amphipathic, and a potential helix capping box occurs at residues α3-8. We introduced truncations, deletions, and mutations into αN1-22 peptide and examined their effects on binding to the δ subunit. The deletions and mutations were introduced also into the N-terminal region of the uncA (α subunit) gene to determine effects on cell growth in vivo and membrane ATP synthase activity in vitro. Effects seen in the peptides were well correlated with those seen in the uncA gene. The results show that, with the possible exception of residues close to the initial Met, all of the αN1-22 sequence is required for binding of δ to α. Within this sequence, an amphipathic helix seems important. Hydrophobic residues on the predicted nonpolar surface are important for δ binding, namely αIle-8, αLeu-11, αIle-12, αIle-16, and αPhe-19. Several or all of these residues probably make direct interaction with helices 1 and 5 of δ. The potential capping box sequence per se appeared less important. Impairment of α/δ binding brings about functional impairment due to reduced level of assembly of ATP synthase in cells.

Original languageEnglish
Pages (from-to)25673-25679
Number of pages7
JournalJournal of Biological Chemistry
Issue number24
StatePublished - Jun 11 2004


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