Stoichiometry of H+ ejection during respiration-dependent accumulation of Ca2+ by rat liver mitochondria.
We have investigated the energy-dependent uptake of Ca2+ by rat liver mitochondria with succinate as respiratory substrate with rotenone added to block NAD-linked electron transport. In the presence of 3-hydroxybutyric or other permeant monocarboxylic acids Ca2+ was taken up to extents approaching those seen in the presence of phosphate. The quantitative relationship between cation and anion uptake was determined from the slope of a plot of 3-hydroxybutyrate uptake against Ca2+ uptake, a method which allowed determination of the stoichiometry without requiring ambiguous corrections for early nonenergized or nonstoichiometric binding events. This procedure showed that 2 molecules of 3-hydroxtbutyrate were accumulated with each Ca2+ ion. Under these conditions close to 2 Ca2+ ions and 4 molecules of 3-hydroxybutyrate were accumulated per pair of electrons per energy-conserving site of the respiratory chain. Since 3-hydroxybutyrate must be protonated to pass the membrane as the undissociated free acid, it is concluded that 4 protons were ejected (and subsequently reabsorbed) per pair of electrons per energy-conserving site, in contrast to the value 2.0 postulated by the chemiosmotic hypothesis.
D000818:Animals; D000968:Antimycin A / Q000494:pharmacology; D001693:Biological Transport, Active; D002118:Calcium / Q000378:metabolism*; D002264:Carboxylic Acids / Q000378:metabolism / Q000494:pharmacology; D004579:Electron Transport; D006859:Hydrogen / Q000378:metabolism*; D006863:Hydrogen-Ion Concentration; D006885:Hydroxybutyrates / Q000378:metabolism; D066298:In Vitro Techniques; D007700:Kinetics; D008297:Male; D008930:Mitochondria, Liver / Q000187:drug effects / Q000378:metabolism*; D010101:Oxygen Consumption / Q000187:drug effects; D051381:Rats; D012402:Rotenone / Q000494:pharmacology; D013386:Succinates / Q000378:metabolism
D002264:Carboxylic Acids; D006885:Hydroxybutyrates; D013386:Succinates; D012402:Rotenone; D000968:Antimycin A; D006859:Hydrogen; D002118:Calcium