The role of the betaDELSEED motif of F1-ATPase: propagation of the inhibitory effect of the epsilon subunit.
In F(1)-ATPase, a rotary motor enzyme, the region of the conserved DELSEED motif in the beta subunit moves and contacts the rotor gamma subunit when the nucleotide fills the catalytic site, and the acidic nature of the motif was previously assumed to play a critical role in rotation. Our previous work, however, disproved the assumption (Hara, K. Y., Noji, H., Bald, D., Yasuda, 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 the epsilon subunit, an intrinsic inhibitor, was unable to inhibit the ATPase activity of a mutant thermophilic F(1)-ATPase in which all of the five acidic residues in the DELSEED motif were replaced with alanines, although the epsilon subunit in the mutant F(1)-ATPase assumed the inhibitory form. In addition, the replacement of basic residues in the C-terminal region of the epsilon subunit by alanines caused a decrease of the inhibitory effect. Partial replacement 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 subunit induced a partial effect. We here conclude that the epsilon subunit exerts its inhibitory effect through the electrostatic interaction with the DELSEED motif of the beta subunit.
Hara
K Y
KY
Chemical Resources Laboratory, R-1, Tokyo Institute of Technology, Nagatsuta 4259, Yokohama, 226-8503, Japan.
Kato-Yamada
Y
Y
Kikuchi
Y
Y
Hisabori
T
T
Yoshida
M
M
D000255:Adenosine Triphosphate / Q000378:metabolism; D020816:Amino Acid Motifs; D004795:Enzyme Stability; D007700:Kinetics; D008954:Models, Biological; D009154:Mutation; D020836:Protein Structure, Quaternary; D017433:Protein Structure, Secondary; D021122:Protein Subunits; D006180:Proton-Translocating ATPases / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D055672:Static Electricity
D021122:Protein Subunits; D000255:Adenosine Triphosphate; D006180:Proton-Translocating ATPases