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G F Azzone

Publications and source records attributed to G F Azzone.

At least 91 records · Page 5Linked to original sources
PubMed1977

The generation of the proton electrochemical potential and its role in energy transduction.

The evidence that all energy transducing membranes can generate a proton electrochemical potential difference, delta micronH, across the membrane and that this potential can be used to transfer energy among energy transducing units and to generate ATP, has increased the interest for the view that delta micronH plays an obligatory role in energy transduction and ATP synthesis. In the present article we shall concentrate on two experimental questions related with the generation and role of delta micronH: (a) the charge/site ratio; (b) the relation between the proton electrochemical potential on one side and the cation electrochemical potential, the phosphate potential and the redox potential on the other. We shall then discuss the view that energy transduction corresponds to a molecular energy machine rather than to a fuel cell.

Adenosine Triphosphate↗
PubMed1977

Proton electrochemical potential in steady state rat liver mitochondria.

Delta approximately muH has been determined in steady state mitochondria by measuring the magnitude of delta pH on the distribution of acetate and of deltapsi on the distribution of K+, tetraphenylphosphonium, Ca2+, Sr2+ and Mn2+. (1) The matrix concentration of divalent cations has been calculated from the total cation uptake, from the increase of matrix volume and from the ESR sextet signal of Mn(H2O)L2+. The [cat2+]i based on osmotic data is about five times higher than that based on ESR measurements. The [cat2+]i based on total uptake is much higher than that based on osmotic data at low cation/protein ratios. (2) In the presence of 10 mM acetate the maximal deltapsi on Ca2+ is about 130 mV and on Sr2+ is 95 mV. Deltapsi on Mn2+ is 91 or 109 mV, according to whether [cat2+-a)i is calculated from ESR or osmotic data. Under the same conditions, deltapH is about 60 mV. Hence delta approximately muH on divalent cations is between 151 and 190 mV. (3) Deltapsi on K+, in valinomycin treated mitochondria with 10 mM acetate or 2 mM Pi, drops from 200 mV, at low [K+]0 to almost zero parallel to the increase of [K+]0. DeltapH is 30 mV at low [K+]0 and about 42 mV at 600 muM K+. Hence delta approximately muH drops from 22m mV lower values with the increase of [K+]0. (4) Maximal deltapsi on triphenylmethylphosphonium is 140 mV. (5) When delta approximately muH is measured simultaneously on divalent cations and on K+, the values on K+ tend to approach those on Ca2+ while those on Sr2+ are about 50 mV lower. (6) It is concluded that the steady state mitochondrial energy potential is equivalent to a delta approximately muH between 150 and approx. 190 mV.

Acetates↗
PubMed1976

Anion and amine uptake and uncoupling in submitochondrial particles.

1. Unlike chloroplasts, submitochondrial particles are not uncoupled by nigericin + KCl or NH4Cl. Also the uncoupling effect of lipophilic anions is largely independent of the addition of weak bases. 2. Low concentrations of permeant anions cause a shift of the steady-state energy level rather than a cycle of energy utilization. The degree of inhibition of ATP synthesis by tetraphenylboron is larger than required for the uptake of the anion. 3. Lipophilic anions such as bromthymolblue, bromcresolpurple, and 8-anilino-1-napthalene sulphonate cause a pH-independent, 50% uncoupling in submitochondrial particles at concentrations of 3, 30 and 30 muM, respectively. The passive interaction of bromthymolblue and bromcresolpurple appears as a pH-dependent distribution between two pHases. ATP causes a pH-independent slight shift in the anion distribution, with negligible anion accumulation. 4. Addition of amines to energized submitochondrial particles results in two types of effects; uptake of amines and uncoupling. While in chloroplasts amine uptake and uncoupling are closely associated, this is not the case in submitochondrial particles. The uncoupling effect is observed only with lipophilic and not with hydrophilic amines, and the degree of uncoupling increases with the lipophilicity of the amines. The amine uptake, on the other hand, is accompanied by negligible uncoupling. 5. While the uptake of amines is dependent on the presence of non-permeant anions, such as Cl-, the uncoupling effect is independent of Cl-. Furthermore the amine uncoupling is markedly enhanced by lipophilic anions. 6. The view is discussed that the uncoupling effect of lipophilic anions and lipophilic amines in submitochondrial particles is due to a catalytic energy dissipation rather than to a stoichiometry energy utilization. The molecular mechanism of uncoupling presumably involves a cycling of charges after a perturbation of the membrane structure.

Amines↗
PubMed1976

Mechanism of active shrinkage in mitochondria. I. Coupling between weak electrolyte fluxes.

1. A passive penetration of (NH4)2 HPO4 or of K2HPO4+nigericin occurs in respiratory-inhibited liver mitochondria. Addition of succinate at the end of the passive swelling initiates a shrinkage phase which leads to restoration of the initial mitochondrial volume. The rate and time of onset of the active shrinkage depend on the degree of stretching of the mitochondrial membrane. The rate of active shrinkage increases proportionally to the concentration of nigericin while it is strongly inhibited by valinomycin. 2. A number of SH inhibitors such as N-ethylmaleimide, p-chloromercuribenzoate, p-chloromercuriphenylsulphonate, dithiobisnitrobenzoate, exert a marked enhancing effect on the rate of shrinkage. The enhancing effect parallels titration of the phosphate carrier and inhibition of the passive phosphate efflux. In contrast, mersalyl is a powerful inhibitor of the rate of active shrinkage. The inhibition parallels that on phosphate passive efflux and requires higher mersalyl concentrations in respect to inhibition of phosphate influx. 3. The active shrinkage is discussed in terms of (a) a mechanoenzyme, (b) an electrogenic proton pump and (c) a proton-driven Pi pump.

Animals↗
PubMed1976

Mechanism of active shrinkage in mitochondria. II. Coupling between strong electrolyte fluxes.

1. Addition of succinate to valinomycin-treated mitochondria incubated in KCL causes a large electrolyte penetration. The process depends on a steady supply of energy and involves a continuous net extrusion of protons. Rates of respiration and of electrolyte penetration proceed in a parallel manner. 2. A passive penetration of K+ salt of permeant anions occurs in respiratory-inhibited mitochondria after addition of valinomycin. Addition of succinate at the end of the passive swelling starts an active extrusion of anions and cations with restoration of the initial volume. The shrinkage is accompanied by a slow reuptake of protons. The initiation of the active shrinkage correlates with the degree of stretching of the inner membrane. The extrusion of electrolytes is inhibited by nigericin, while it is only slightly sensitive to variations of the valinomycin concentration larger than two orders of magnitude. 3. Passive swelling and active shrinkage occurs also when K+ is replaced by a large variety of organic cations. The rate of organic cation penetration is enhanced by tetraphenylboron, while the rate of electrolyte extrusion is insensitive to variation of the tetraphenylboron concentration. 4. Active shrinkage, either with K+ or organic cation salts, is inhibited by weak acids. The phosphate inhibition is removed by SH inhibitors. The active shrinkage is also inhibited by mersalyl to an extent of about 60%. 5. Three models of active shrinkage are discussed: (a) mechanoprotein, (b) electrogenic proton pump, and (c) proton-driven cation anion pump.

Animals↗
PubMed1976

The effect of endogenous phosphate on the H+/Mn2+ ratio and the state of Mn2+ in the mitochondrial matrix.

1. Kinetics and stoichiometry of H+ extrusion and reuptake and of Mn2+ uptake and release have been measured in respiring liver mitochondria in the absence of external added Pi. H+ and Mn2+ fluxes are parallel during aerobic cation uptake but not during uncoupler induced cation release. The H+/Mn2+ is 1.24. Addition of SH reagents, in concentrations inhibiting the Pi carrier, modifies the kinetics of H+ extrusion and of Mn2+ uptake and release. The slow phase of uncoupler induced Mn2+ release is diminished. The H+/Mn2+ is increased to 1.72. Addition of SH reagents, after the phase of aerobic uptake is completed, results in a significant reduction of the extent of uncoupler-induced Mn2+ release. The extent of reuptake of endogenous Pi during aerobic uptake of Mn2+ is about 8 nmol x mg protein-1. 2. Aerobic uptake of Mn2+ in the absence of external Pi results in an electron spin resonance spectrum which is the sum of two components. One, denoted as S, corresponds to Mn(H2O)2+(6). Another denoted as E, reflects spin exchange narrowing. In contrast to previous claims the following evidence suggests that the spin exchange component is due to Mn3(PO4)2 precipitate: (a) the dimension of the spin exchange spectrum is markedly reduced by abolition of Pi transport; (b) the spin exchange spectrum is released very slowly by addition of uncouplers under conditions where uncouplers cause a rapid deenergization of mitochondria, reuptake of H+ and release of cations; (c) the free matrix Mn2+ is released slowly after addition of uncoupler if there is a large spin exchange signal; howeover the free matrix Mn2+ is abolished rapidly by uncoupler when formation of the spin exchange signal is prevented by pretreatment with Ca2+; (d) the band width of the spin exchange fraction is independent of the Mn2+/protein ratio either under kinetic or steady state conditions; (e) the experimental spectrum recalls closely that obtained by computer simulation by assuming it as a combination of Mn(H2O)2+(6) and Mn3(PO4)2. 3. It is concluded that endogenous Pi affects the process of aerobic divalent cation uptake. A part of Mn2+ uptake in the absence of externally added anions, consists of a Mn3(PO4)2 precipitate. This accounts for a H+/Mn2+ ratio lower than 2.

Animals↗