The c-ring ion binding site of the ATP synthase from Bacillus pseudofirmus OF4 is adapted to alkaliphilic lifestyle.
In the c-ring rotor of ATP synthases ions are shuttled across the membrane during ATP synthesis by a unique rotary mechanism. We investigated characteristics of the c-ring from the alkaliphile Bacillus pseudofirmus OF4 with respect to evolutionary adaptations to operate with protons at high environmental pH. The X-ray structures of the wild-type c13 ring at pH 9.0 and a 'neutralophile-like' mutant (P51A) at pH 4.4, at 2.4 and 2.8 Å resolution, respectively, reveal a dependency of the conformation and protonation state of the proton-binding glutamate (E(54) ) on environmental hydrophobicity. Faster labelling kinetics with the inhibitor dicyclohexylcarbodiimide (DCCD) demonstrate a greater flexibility of E(54) in the mutant due to reduced water occupancy within the H(+) binding site. A second 'neutralophile-like' mutant (V21N) shows reduced growth at high pH, which is explained by restricted conformational freedom of the mutant's E(54) carboxylate. The study directly connects subtle structural adaptations of the c-ring ion binding site to in vivo effects of alkaliphile cell physiology.
Department of Structural Biology, Max Planck Institute of Biophysics, Max-von-Laue-Str. 3, 60438, Frankfurt am Main, Germany.
Laura
L
Preiss
Julian D
JD
Langer
David B
DB
Hicks
Jun
J
Liu
Ozkan
O
Yildiz
Terry A
TA
Krulwich
Thomas
T
Meier
D001407:Bacillus / Q000201:enzymology*; D025243:Bacterial Proton-Translocating ATPases / Q000037:antagonists & inhibitors / Q000737:chemistry* / Q000378:metabolism*; D001665:Binding Sites; D018360:Crystallography, X-Ray; D004024:Dicyclohexylcarbodiimide / Q000494:pharmacology; D006863:Hydrogen-Ion Concentration
D004024:Dicyclohexylcarbodiimide; D025243:Bacterial Proton-Translocating ATPases