The role of the beta DELSEED motif of F-1-ATPase - Propagation of the inhibitory effect of the epsilon subunit

In F-1-ATPase, a rotary motor enzyme, the region of the conserved DELSEED m otif in the beta subunit moves and contacts the rotor gamma subunit when th e nucleotide fills the catalytic site, and the acidic nature of the motif w as previously assumed to play a critical role in rotation. Our previous wor k, however, disproved the assumption (Hara, K. Y., Noji, H., Bald, D., Yasu da, R., Kinosita, K., Jr., and Yoshida, M. (2000) J. Biol. Chem. 275, 14260 -14263), and the role of this motif remained unknown. Here, we found that t he E subunit, an intrinsic inhibitor, was unable to inhibit the ATPase acti vity of a mutant thermophilic F-1-ATPase in which all of the five acidic re sidues in the DELSEED motif were replaced with alanines, although the epsil on subunit in the mutant F-1-ATPase assumed the inhibitory form. In additio n, the replacement of basic residues in the C-terminal region of the epsilo n subunit by alanines caused a decrease of the inhibitory effect. Partial r eplacement of the acidic residues in the DELSEED motif of the beta subunit or of the basic residues in the C-terminal alpha -helix of the epsilon subu nit induced a partial effect. We here conclude that the epsilon subunit exe rts its inhibitory effect through the electrostatic interaction with the DE LSEED motif of the beta subunit.