Subunit rotation of ATP synthase embedded in membranes: a or beta subunit rotation relative to the c subunit ring.

Abstract

ATP synthase F(o)F(1) (alpha(3)beta(3)gammadelta epsilon ab(2)c(10-14)) couples an electrochemical proton gradient and a chemical reaction through the rotation of its subunit assembly. In this study, we engineered F(o)F(1) to examine the rotation of the catalytic F(1) beta or membrane sector F(o) a subunit when the F(o) c subunit ring was immobilized; a biotin-tag was introduced onto the beta or a subunit, and a His-tag onto the c subunit ring. Membrane fragments were obtained from Escherichia coli cells carrying the recombinant plasmid for the engineered F(o)F(1) and were immobilized on a glass surface. An actin filament connected to the beta or a subunit rotated counterclockwise on the addition of ATP, and generated essentially the same torque as one connected to the c ring of F(o)F(1) immobilized through a His-tag linked to the alpha or beta subunit. These results established that the gamma epsilon c(10-14) and alpha(3)beta(3)deltaab(2) complexes are mechanical units of the membrane-embedded enzyme involved in rotational catalysis.