Open AccessClarification of Factors Influencing the Nature and Magnitude of the Protonmotive Force in Bovine Heart Submitochondrial Particles
Author(s) -
BRANCA Donata,
FERGUSON Stuart J.,
SORGATO M. Catia
Publication year - 1981
Publication title -
european journal of biochemistry
Language(s) - English
Resource type - Journals
eISSN - 1432-1033
pISSN - 0014-2956
DOI - 10.1111/j.1432-1033.1981.tb05340.x
Subject(s) - submitochondrial particle , tris , chemistry , hepes , atpase , membrane potential , electrochemical gradient , biophysics , substrate (aquarium) , chemiosmosis , phosphate , sucrose , membrane , osmotic concentration , stereochemistry , atp synthase , biochemistry , enzyme , biology , ecology
The magnitude of the protonmotive force, and its division between pH gradient and membrane potential components has been further characterised in submitochondrial particles. In a reaction medium containing sucrose for osmotic support and 4‐(2‐hydroxyethyl)‐1‐piperazineethanesulfonate (Hepes) as buffer, with succinate as substrate, the total protonmotive force reached a maximum value of 245 mV. The presence of Cl − enhanced the pH gradient with a partial but not fully compensating decrease in the membrane potential. When submitochondriaf particles were suspended in a medium of low osmolarity consisting of phosphoric acid neutralised with Tris. again with succinate as substrate, the protonmotive force was lower and did not exceed 185 mV, and the pH gradient component was equivalent to 25 mV or less. The final phosphorylation potential, ΔG p ,maintained by the particles was higher in the phosphate/Tris medium (46–47.7 kJ mol −1 ) than in the sucrose/Hepes/KCl medium (43.7 kJ mol −1 ). Thus, comparison of the phosphorylation potential with the protonmotive force would suggest that the mechanistic stoichiometry H + /ATP (H + translocated per molecule of ATP synthesied) for the ATPase enzyme is 3 in the former medium and 2 in the latter, which might be taken to indicate two different types of mechanists required for ATP synthesis. However it is questioned whether a comparison of the protonmotive force with ΔG p in terms of equilibrium thermodynamics ought not to be complemented by analysis in ierms of linear non‐equilibrium thermodynamics. The latter treatment shows that it is possible to estimate only a value for the product of a phenomenological stoichiometry and the degree of coupling, which can be variable, but not the mechanistic stoichiometry. This treatment can also rationalise the observation of the higher ΔG p in reaction conditions where the lower values for Δp are estimated. Irrespective of possible explanations, the data show flow an unprejudiced choice of reaction conditions can lead to different conclusions about the relationship between the phosphorylation potential and the protonmotive force.