Valinomycin diffusion potential

K+ ions in the presence of valinomycin do not distribute passively at electrochemical equilibrium rather, this represents a nonequilibrium state in which creates a diffusion potential following which protons move. 

The addition of valinomycin, a K+ ionophore, leads to generation of a diffusion potential through efflux of K+ down its concentration gradient. The F protein permits uptake of H+ by the vesicles (which are impermeable to H+ in the absence of F ) in response to this diffusion-generated gradient. 

The electrochromic shift of the carotenoids is usually calibrated with K-diffusion potential in the presence of valinomycin. One problem is that the shifts observed in respiring chromatophores (where the proton electrochemical potential is predominantly in the form of a membrane potential) are much larger than those induced by the calibrating diffusion potential, so that an extensive extrapolation is required. Thus, the carotenoids in illuminated chromatophores may indicate a membrane potential in excess of 300 mV, whereas the distribution of CNS, an electrically permeant anion, in the same system only indicates 140 mV [32]. The extent of this discrepancy, and the uncertainty as to whether the carotenoids see the bulk-phase potential, or only the local electrical field within the membrane, limits the confidence with which carotenoids may be used for quantitative as opposed to qualitative potential measurements. 

Membrane Potential produced b Diffusion Potential. In 1973,Uribe and reported that addition of valinomycin and to chloroplasts enhanced ATP synthesis elicited by acid-base transition and attributed... 

In a typical experiment, proton flux might be driven by placing liposomes that contain pH 8 buffer in a solution buffered at pH 6, so that the initial proton flux is driven by 10 6-M protons outside and 10 6 hydroxide ions inside. Buffers are required to make the decay rate of pH gradients sufficiently slow so that initial rates can be conveniently estimated. It is also necessary to release proton diffusion potentials by addition of valinomycin or a permeant anion otherwise the proton flux will be limited by counterion flux (9). 

Figure 4. The dependence of the ellipticity of bacteriorhodopsin reconstituted in vesicles on the electric diffusion potentials. The ordinate is the decrease of the CD signal at 210 nm induced by 10 7-M valinomycin. The abscissa is the electrical potentials calculated by the Nemst equation. The sign indicates the polarity inside the vesicle.

Fig. 1 shows that the curves obtained with 0.5 and 5 mM K in the internal volume were displaced by only about 0.35 pH-units, or approx. 21 mV from one another. This is due to the effect of the membrane electrical capacitance on the distribution of at equilibrium [4,7]. We used the method of Apell and Bersch [ ] to calculate equilibrium values of the -diffusion potential after addition of proteoliposomes with a known internal -concentration to a medium with 50 mM K, in the presence of valinomycin. Fig. 2 shows the dependence of these diffusion potentials on the internal diameter of the proteoliposomes. The dashed line in Fig. 2 shows that with proteoliposomes of 27 nm internal diameter, the -diffusion potential obtained with an initial internal K -concentration of 0.5 mM is only 21 mV higher than the one obtained with an initial internal -concentration of 5 mM. The diffusion potential obtained in the latter case is 52 mV. These diffusion potentials correspond with ApH-values of 0.36 and 0.88 units, respectively. This is in good agreement with the results shown in Fig. 1, and the required internal diameter of 27 nm is in good agreement with electron-microscopic and other evidence on the size of the proteoliposomes [2]. Furthermore, Fig. 2 shows that vesicles of this diameter generate a K -diffusion potential of only 77 mV even if the initial internal -concentration is zero. Since ATP-synthesis was observed only above a threshold Apjj+ of 90 mV (Fig. 1), this explains why... 

A series of experiments was first carried out to determine the response of the carotenoid band shift to K+-diffusion potentials. The procedure was similar to that employed on earlier occasions (Jackson, Crofts, 1969 Clark, Jackson, 1981) except that sodium ferri- and ferrocyanide were present as a redox buffer. The treatment of the chromatophores with valinomycin was followed by a period of 10 min to allow ionic equilibration across the membrane. The extent of the absorbance change corresponding to the carotenoid band shift, resulting from a subsequent KCl addition was plotted as a function of the final KCl concentration as shown in Fig.3. 

A comparison of this change to a change brought about by potassium diffusion potentials in the presence of valinomycin indicated that potentials in excess of lOOmV could be generated by light activation. 

Another analytically useful phenomenon in electrolysis at ITIES is ion transfer faciUtated by ionophores present in the non-aqueous phase [8]. If the ionophore is present at a low concentration in the non-aqueous phase and the aqueous phase contains a large concentration of the cation that is bound in a complex with the ionophore (for example as a component of the base electrolyte), then a voltammetric wave controlled by diffusion of the ionophore toward the ITIES or by diffusion of the complex formed away from the ITIES into the bulk of the organic phase appears at a potential lower than the potential of simple cation transfer. The peak height of this wave is proportional to the ionophore concentration in the solution and can be used for the determination (fig. 9.8). This effect has been observed with valinomycin, nonactin, cycUc polyethers and other substances [2,3,23]. The half-wave potential of these waves is... 

The potential difference across the floating lipid bilayer on top of the thiolspacer SAM (i.e. the transmembrane potential) can be estimated by noting that its zero value is practically attained at the applied potential E at which the absolute potential difference across the whole interphase, A (f/SCE-t-0.250 V) (see Eq. 9), is equal to the calculated value of Xt. In fact, from Eq. (21) it is apparent that, under these conditions the charge density q = ctm + aoi experienced by the diffuse-layer ions is zero, just as the electric field across the floating lipid bdayer. The applied potential at which the transmembrane potential equals zero can also be approximately estimated by incorporating valinomycin in the lipid bilayer [37, 121]. Valinomycin is a K+-selective ion carrier that may shuttle K+ ions across... 

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