![]() ![]() Thus, up to 240 protons per glucose are translocated across the membrane. This involves vectorial translocation of up to 120 protons by the respiratory chain, which will be taken up again by the ATPase. For example, a bacterium respiring a single molecule of glucose with oxygen gains about 30 ATP ( Rich, 2003). As the ATPase takes up 3–4 H + per ATP ( Cross and Müller, 2004 Steigmüller et al., 2008 Petersen et al., 2012), which before have been pumped out driven by respiratory or photosynthetic electron transport, proton translocation across a membrane is the most common process in living cells. ![]() The pmf is then used to drive ATP conservation by the membrane-bound ATP synthase (or simply ATPase), which is also an electrogenic proton transport system. The most important electrogenic vectorial processes are proton translocation during respiration and photosynthesis, which build up the proton-motive force (pmf, composed of the trans-membrane proton gradient, and membrane potential). By contrast, electrogenic transport (e.g., symport of 2 H + per anion A –) affects the electrical potential across the membrane. Some vectorial processes – named electroneutral – are not going along with a net charge transfer (e.g., transport of an anion A – in symport with a single proton H +). The resulting pH effects are transient and compensated by counteracting cyclic transport processes. By far more common are vectorial processes which translocate protons or metabolites across a membrane without changing their overall amount. As such reactions change the amount of protons in the system they are named scalar. First, production or consumption of acidic or basic metabolites results in permanent pH effects. Our approach allows for studying a broad variety of proton-related metabolic activities at micromolar concentrations and time scales of seconds to minutes.Īll living cells, mitochondria, chloroplasts or bacteria cause various types of pH changes. We could verify the kinetic simulation parameters found with proton.exe using series of increasing additions of the reactants. This method gave lower but more realistic values than logarithmic extrapolation. H +/e – ratios of electron-transport driven proton translocation were calculated by simulation with proton.exe. To study electron-transport driven proton translocation, the membrane potential was neutralized by addition of KSCN (100 mM). For reductant pulse experiments, small amounts of H 2-saturated KCl were added to cells incubated under N 2 with an excess of one of the above-mentioned electron acceptors. Two types of experiments were performed: In oxidant pulse experiments, cells were kept under H 2, and micromolar amounts of sulfate, nitrate or nitrite were added. We analysed sulfate uptake by proton-sulfate symport, scalar alkalinization by sulfate reduction to sulfide, as well as nitrate and nitrite reduction to ammonia, and electron transport-coupled proton translocation. In our experiments, the versatile sulfate-reducing bacterium Desulfovibrio desulfuricans CSN (DSM 9104) was used as model organism. The simulation includes changes of the transmembrane ΔpH, membrane potential and ATP gains, and demonstrates the principles of chemiosmotic energy conservation. The program applies Michaelis-Menten or first-order kinetics to the metabolic processes and allows for parametrization of simultaneously ongoing processes. ![]() Recorded data were used for simulating substrate turnover rates by means of a new freeware app ( proton.exe). ![]() Proton release and uptake induced by metabolic activities were measured in non-buffered cell suspensions by means of a pH electrode. Institute for Chemistry and Biology of the Marine Environment, Carl-von-Ossietzky University of Oldenburg, Oldenburg, Germany. ![]()
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