Rotation of a γ-ε Subunit Domain in the Escherichia coli F1F0-ATP Synthase Complex

Aggeler, Robert; Ogilvie, Isla; Capaldi, Roderick A.

doi:10.1074/jbc.272.31.19621

Cited by 110 publications

(52 citation statements)

References 36 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…Cross-linking studies (Aggeler et al 1997) have indicated that e is probably a part of the rotor and d belongs to the stator. When an actin ¢lament was bound to e instead of g, the ¢lament also rotated counterclockwise, although the rotary speed was somewhat lower (KatoYamada et al 1998).…”

Section: (D) F O Awaits More Experimentsmentioning

confidence: 99%

A rotary molecular motor that can work at near 100% efficiency

Kinosita

Yasuda

Noji

et al. 2000

Phil. Trans. R. Soc. Lond. B

225

141

View full text Add to dashboard Cite

A single molecule of F 1 -ATPase is by itself a rotary motor in which a central g-subunit rotates against a surrounding cylinder made of a 3 b 3 -subunits. Driven by the three bs that sequentially hydrolyse ATP, the motor rotates in discrete 1208 steps, as demonstrated in video images of the movement of an actin ¢lament bound, as a marker, to the central g-subunit. Over a broad range of load (hydrodynamic friction against the rotating actin ¢lament) and speed, the F 1 motor produces a constant torque of ca. 40 pN nm. The work done in a 1208 step, or the work per ATP molecule, is thus ca. 80 pN nm. In cells, the free energy of ATP hydrolysis is ca. 90 pN nm per ATP molecule, suggesting that the F 1 motor can work at near 100% e¤ciency. We con¢rmed in vitro that F 1 indeed does ca. 80 pN nm of work under the condition where the free energy per ATP is 90 pN nm. The high e¤ciency may be related to the fully reversible nature of the F 1 motor: the ATP synthase, of which F 1 is a part, is considered to synthesize ATP from ADP and phosphate by reverse rotation of the F 1 motor. Possible mechanisms of F 1 rotation are discussed.

show abstract

Section: (D) F O Awaits More Experimentsmentioning

confidence: 99%

A rotary molecular motor that can work at near 100% efficiency

Kinosita

Yasuda

Noji

et al. 2000

Phil. Trans. R. Soc. Lond. B

225

141

View full text Add to dashboard Cite

show abstract

“…Recent evidence strongly suggests that ␥ and probably ⑀ rotate relative to ␣ and ␤ during catalysis (13)(14)(15)(16). Several studies have implied that the ␦ subunit is located near the top of the ␣␤ cluster (17)(18)(19)(20).…”

mentioning

confidence: 99%

The b and δ Subunits of the Escherichia coli ATP Synthase Interact via Residues in their C-terminal Regions

McLachlin¹,

Bestard

Dunn

1998

Journal of Biological Chemistry

View full text Add to dashboard Cite

An affinity resin for the F 1 sector of the Escherichia coli ATP synthase was prepared by coupling the b subunit to a solid support through a unique cysteine residue in the N-terminal leader. b 24 -156 , a form of b lacking the N-terminal transmembrane domain, was able to compete with the affinity resin for binding of F 1 . Truncated forms of b 24 -156 , in which one or four residues from the C terminus were removed, competed poorly for F 1 binding, suggesting that these residues play an important role in b-F 1 interactions. Sedimentation velocity analytical ultracentrifugation revealed that removal of these C-terminal residues from b 24 -156 resulted in a disruption of its association with the purified ␦ subunit of the enzyme. To determine whether these residues interact directly with ␦, cysteine residues were introduced at various C-terminal positions of b and modified with the heterobifunctional cross-linker benzophenone-4-maleimide. Cross-links between b and ␦ were obtained when the reagent was incorporated at positions 155 and 158 (two residues beyond the normal C terminus) in both the reconstituted b 24 -156 -F 1 complex and the membranebound F 1 F 0 complex. CNBr digestion followed by peptide sequencing showed the site of cross-linking within the 177-residue ␦ subunit to be C-terminal to residue 148, possibly at Met-158. These results indicate that the b and ␦ subunits interact via their C-terminal regions and that this interaction is instrumental in the binding of the F 1 sector to the b subunit of F 0 .In the process of oxidative phosphorylation or photophosphorylation, the electron transport chain generates a transmembrane proton gradient. The ATP synthase, or F 1 F 0 -ATPase, allows protons to flow down this electrochemical gradient and uses the energy obtained to synthesize ATP (for reviews, see Refs. 1-4). Under appropriate conditions ATP synthase can hydrolyze ATP to pump protons. The enzyme is composed of two sectors; the F 0 sector is membrane-integral and is responsible for proton translocation, and the F 1 sector is attached to the membrane via F 0 and houses the catalytic sites for ATP synthesis. F 1 is easily detached from the membrane and can be purified as a soluble protein with ATPase activity.ATP synthases contain at least eight types of subunits. In the relatively simple enzyme from Escherichia coli, the F 1 sector has the stoichiometry ␣ 3 ␤ 3 ␥ 1 ␦ 1 ⑀ 1 , whereas F 0 is composed of three subunits of stoichiometry a 1 b 2 c 9 -12 . The a and c subunits, but not b, contain residues essential for the translocation of protons across the membrane (3). The 156-residue b subunit is believed to span the membrane once at its hydrophobic N terminus, whereas the remainder of the protein is very hydrophilic. b is thought to exist as a dimer in the complex (5-7), and proteolysis studies have shown that the hydrophilic region of b is required for the association of F 1 with the membrane (7-9). Removal of two residues from the C terminus of b disrupts normal assembly of the complex (10), as does mut...

show abstract

“…E. coli strains used were XL1-Blue (Stratagene) for site-directed mutagenesis and for cloning and the unc Ϫ RA1 (29) to express the ATP synthase. Plasmids used were pRA13 (30) containing the wild-type ␤, ␥, and genes and pRA197 (31) containing the wild-type unc operon except that two c subunits were fused through an additional sequence, and a cysteine was introduced in position 42 in the latter of the two c subunits (ccЈ).…”

Section: Methodsmentioning

confidence: 99%

Rotation of the c subunit oligomer in fully functional F1Fo ATP synthase

Tsunoda¹

2001

Proceedings of the National Academy of Sciences

View full text Add to dashboard Cite

The F1Fo-type ATP synthase is the smallest motor enzyme known. Previous studies had established that the central ␥ and subunits of the F 1 part rotate relative to a stator of ␣3␤3 and ␦ subunits during catalysis. We now show that the ring of c subunits in the Fo part moves along with the ␥ and subunits. This was demonstrated by linking the three rotor subunits with disulfide bridges between cysteine residues introduced genetically at the interfaces between the ␥, , and c subunits. Essentially complete cross-linking of the ␥, , and c subunits was achieved by using CuCl2 to induce oxidation. This fixing of the three subunits together had no significant effect on ATP hydrolysis, proton translocation, or ATP synthesis, and each of these functions retained inhibitor sensitivity. These results unequivocally place the c subunit oligomer in the rotor part of this molecular machine.

show abstract

Rotation of a γ-ε Subunit Domain in the Escherichia coli F1F0-ATP Synthase Complex

Cited by 110 publications

References 36 publications

A rotary molecular motor that can work at near 100% efficiency

A rotary molecular motor that can work at near 100% efficiency

The b and δ Subunits of the Escherichia coli ATP Synthase Interact via Residues in their C-terminal Regions

Rotation of the c subunit oligomer in fully functional F1Fo ATP synthase

Contact Info

Product

Resources

About