Subunit rotation of ATP synthase embedded in membranes: a or beta subunit rotation relative to the c subunit ring.
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.
D000199:Actins / Q000737:chemistry / Q000378:metabolism; D000255:Adenosine Triphosphate / Q000378:metabolism; D004024:Dicyclohexylcarbodiimide / Q000494:pharmacology; D004926:Escherichia coli / Q000201:enzymology / Q000235:genetics; D029968:Escherichia coli Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D008566:Membranes / Q000201:enzymology; D008567:Membranes, Artificial; D008854:Microscopy, Electron; D008958:Models, Molecular; D015202:Protein Engineering; D021122:Protein Subunits; D006180:Proton-Translocating ATPases / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D012399:Rotation
D000199:Actins; D029968:Escherichia coli Proteins; D008567:Membranes, Artificial; D021122:Protein Subunits; D011994:Recombinant Proteins; D004024:Dicyclohexylcarbodiimide; D000255:Adenosine Triphosphate; D006180:Proton-Translocating ATPases