Comparison of the H+/ATP ratios of the H+-ATP synthases from yeast and from chloroplast.
F(0)F(1)-ATP synthases use the free energy derived from a transmembrane proton transport to synthesize ATP from ADP and inorganic phosphate. The number of protons translocated per ATP (H(+)/ATP ratio) is an important parameter for the mechanism of the enzyme and for energy transduction in cells. Current models of rotational catalysis predict that the H(+)/ATP ratio is identical to the stoichiometric ratio of c-subunits to β-subunits. We measured in parallel the H(+)/ATP ratios at equilibrium of purified F(0)F(1)s from yeast mitochondria (c/β = 3.3) and from spinach chloroplasts (c/β = 4.7). The isolated enzymes were reconstituted into liposomes and, after energization of the proteoliposomes with acid-base transitions, the initial rates of ATP synthesis and hydrolysis were measured as a function of ΔpH. The equilibrium ΔpH was obtained by interpolation, and from its dependency on the stoichiometric ratio, [ATP]/([ADP]·[P(i)]), finally the thermodynamic H(+)/ATP ratios were obtained: 2.9 ± 0.2 for the mitochondrial enzyme and 3.9 ± 0.3 for the chloroplast enzyme. The data show that the thermodynamic H(+)/ATP ratio depends on the stoichiometry of the c-subunit, although it is not identical to the c/β ratio.
Department of Biochemistry and Molecular Biology,  Monash University, Clayton, Victoria 3800,  Australia.
Jan
J
Petersen
Kathrin
K
Förster
Paola
P
Turina
Peter
P
Gräber
D025181:ATP Synthetase Complexes / Q000737:chemistry*; D000255:Adenosine Triphosphate / Q000737:chemistry*; D002021:Buffers; D002138:Calibration; D002384:Catalysis; D002736:Chloroplasts / Q000378:metabolism; D004798:Enzymes / Q000737:chemistry; D006863:Hydrogen-Ion Concentration; D006868:Hydrolysis; D008928:Mitochondria / Q000201:enzymology; D008954:Models, Biological; D018892:Proton-Motive Force; D011522:Protons; D015203:Reproducibility of Results; D012441:Saccharomyces cerevisiae / Q000201:enzymology; D018724:Spinacia oleracea / Q000201:enzymology; D013816:Thermodynamics
D002021:Buffers; D004798:Enzymes; D011522:Protons; D000255:Adenosine Triphosphate; D025181:ATP Synthetase Complexes