Engineering rotor ring stoichiometries in the ATP synthase.
ATP synthase membrane rotors consist of a ring of c-subunits whose stoichiometry is constant for a given species but variable across different ones. We investigated the importance of c/c-subunit contacts by site-directed mutagenesis of a conserved stretch of glycines (GxGxGxGxG) in a bacterial c(11) ring. Structural and biochemical studies show a direct, specific influence on the c-subunit stoichiometry, revealing c(<11), c(12), c(13), c(14), and c(>14) rings. Molecular dynamics simulations rationalize this effect in terms of the energetics and geometry of the c-subunit interfaces. Quantitative data from a spectroscopic interaction study demonstrate that the complex assembly is independent of the c-ring size. Real-time ATP synthesis experiments in proteoliposomes show the mutant enzyme, harboring the larger c(12) instead of c(11), is functional at lower ion motive force. The high degree of compliance in the architecture of the ATP synthase rotor offers a rationale for the natural diversity of c-ring stoichiometries, which likely reflect adaptations to specific bioenergetic demands. These results provide the basis for bioengineering ATP synthases with customized ion-to-ATP ratios, by sequence modifications.
Department of Structural Biology, Max-Planck Institute of Biophysics, 60438 Frankfurt am Main, Germany. denys.pogoryelov@biophys.mpg.de
Denys
D
Pogoryelov
Adriana L
AL
Klyszejko
Ganna O
GO
Krasnoselska
Eva-Maria
EM
Heller
Vanessa
V
Leone
Julian D
JD
Langer
Janet
J
Vonck
Daniel J
DJ
Müller
José D
JD
Faraldo-Gómez
Thomas
T
Meier
D025181:ATP Synthetase Complexes / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D000255:Adenosine Triphosphate / Q000096:biosynthesis; D004591:Electrophoresis, Polyacrylamide Gel; D018625:Microscopy, Atomic Force; D008854:Microscopy, Electron; D008958:Models, Molecular; D009154:Mutation; D011487:Protein Conformation; D011510:Proteolipids / Q000378:metabolism; D020349:Surface Plasmon Resonance
D011510:Proteolipids; C020485:proteoliposomes; D000255:Adenosine Triphosphate; D025181:ATP Synthetase Complexes