Functional domains of epsilon subunit of Escherichia coli H+-ATPase (F0F1).

Mutants of the uncC gene for the epsilon subunit (138 amino acid residues) of Escherichia coli H+-ATPase were isolated: strain KF53 (Gln-72----end) and KF148(SD-) (two base substitutions in the Shine-Dalgarno sequence, GGAGG----AAAGG). These strains did not have F1 bound to membranes and were unable to grow by oxidative phosphorylation. A series of plasmids carrying truncated uncC genes were constructed and introduced into strain KF148(SD-). Analyses of KF148(SD-) cells with different plasmids indicated that the amino-terminal fragment of the epsilon subunit of 78-80 amino acid residues was capable of forming active membrane-bound F1-ATPase, whereas that of 73 residues was not, indicating that the carboxyl-terminal half of the epsilon subunit is not necessary for the active enzyme. Furthermore, results indicated that residues between 73 and 78-80 may have a critical role(s) in binding F1 to F0. Truncated epsilon subunits of 80 and 93 residues were identified in purified F1 from cells carrying the respective uncC genes, and only the latter subunit had intrinsic activity to inhibit ATPase of F1, suggesting that residues between 80 and 93 are essential for the inhibitory activity.