Instrumentation and procedures for real time measurements of proton motive force, membrane potential, Δ<i>p</i>H, proton extrusion, and oxygen uptake in respiring cells and vesicles

Hendler, Richard W.; Setty, Oruganti H.; Shrager, Richard I.; Songco, David C.; Friauf, Walter S.

doi:10.1063/1.1137326

Cited by 16 publications

(4 citation statements)

References 14 publications

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“…It became possible to FIGURE I Fitting of 02 electrode data. At zero time a photographic shutter was opened to initiate respiration in a mitochondrial suspension (18 mg protein in 6 ml) containing 200 mM sucrose,l.5 mM HEPES, 50 mM KCI, 2 ;tM rotenone, 18 ,tg oligomycin, 90 tsg NEM, 2.2 ng valinomycin, 5 mM succinate and 60 to 120 ngA 0, under an atmosphere of 100% carbon monoxide. Data was collected at the rate of 100 electrode readings per s. Top panel: The raw data are shown as a solid line.…”

Section: Discussionmentioning

confidence: 99%

Direct measurement of the initial proton extrusion to oxygen uptake ratio accompanying succinate oxidation by rat liver mitochondria

Setty¹,

Shrager²,

Bunow³

et al. 1986

Biophysical Journal

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The problem of obtaining very early ratios for the H+/O stoichiometry accompanying succinate oxidation by rat liver mitochondria was attacked using new techniques for direct measurement rather than extrapolations based on data obtained after mixing and the recovery of the electrode from initial injection of O2. Respiration was quickly initiated in a thoroughly mixed O2-containing suspension of mitochondria under a CO atmosphere by photolysis of the CO-cytochrome c oxidase complex-. Fast responding O2 and pH electrodes were used to collect data every 10 ms. The response time for each electrode was experimentally measured in each experiment and suitable corrections for electrode relaxations were made. With uncorrected data obtained after 0.8 s, the extrapolation back to zero time on the basis of single-exponential curve fitting confirmed values close to 8.0 as previously reported (Costa et al., 1984). The data directly obtained, however, indicate an initial burst in H+/O ratio that peaked to values of approximately 20 to 30 prior to 50 ms and which was no longer evident after 0.3 s. Newer information and considerations that place all extrapolation methods in question are discussed.

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Section: Discussionmentioning

confidence: 99%

Direct measurement of the initial proton extrusion to oxygen uptake ratio accompanying succinate oxidation by rat liver mitochondria

Setty¹,

Shrager²,

Bunow³

et al. 1986

Biophysical Journal

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“…3) caused a profound membrane-potential decrease, also suggesting a significant synergism between PGLa and magainin-2 in this action. Changes in the reading of the Ph,P+ ion-selective electrode can be converted into apparent membrane potential changes (Hendler et al, 1983). We estimated (data not shown) that the second addition of magainin-2, in the magainin-2 titration (Fig.…”

Section: Synergism Between Pgla and Magainin-2 In Inhibiting Bacteriamentioning

confidence: 96%

Functional Synergism of the Magainins PGLa and Magainin-2 in Escherichia coli, Tumor Cells and Liposomes

Westerhoff¹,

Zasloff²,

Rosner³

et al. 1995

Eur J Biochem

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129

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General rightsIt is not permitted to download or to forward/distribute the text or part of it without the consent of the author(s) and/or copyright holder(s), other than for strictly personal, individual use, unless the work is under an open content license (like Creative Commons). Disclaimer/Complaints regulationsIf you believe that digital publication of certain material infringes any of your rights or (privacy) interests, please let the Library know, stating your reasons. In case of a legitimate complaint, the Library will make the material inaccessible and/or remove it from the website. Please Ask the Library: http://uba.uva.nl/en/contact, or a letter to: Library of the University of Amsterdam, Secretariat, Singel 425, 1012 WP Amsterdam, The Netherlands. You will be contacted as soon as possible. Download date: 10 May 2018Eur. J. Biochem. 228, 257-264 (1995) 0 FEBS 1995 Xenopus laevis skin secretion contains a mixture of magainins, which are small positively charged oligopeptides with antimicrobial activity. In this study, we show that two of these peptides, i.e. magainin-2 and PGLa, are much more active in biological functions when added together than when added alone. This synergy applies for the antimicrobial activity of these peptides, and for the toxic effects on tumor cells. We show that this peptide combination is also synergistic when permeabilizing protein-free liposomes for glucose, when dissipating the membrane potential in cytochrome oxidase liposomes and Escherichia coli, and, reversibly, when stimulating respiration in the liposomes. The occurrence of synergy in these diverse systems (complex and simple) suggests that the biological synergy results from synergy in the primary activity of the magainin peptides, namely the permeabilization of free-energy transducing membranes, possibly by forming a multimeric transmembrane pore of mixed peptide composition. The antimicrobial activity of X. laevis skin secretions may be greatly enhanced by the application of this binary weapon.

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“…The cell and all of the probes were placed in a grounded metal Faraday cage. The signals from all electrodes were passed to a custom electronic interface which was able to provide gains, speed the response of the pH electrode, reduce noise by common mode rejection between grounded shields on the pH and reference electrodes, and electronic filters (Hendler et al, 1983;Setty et al, 1986). The optics signal was passed through an Oriel Corp. #53870 interference filter with a sharp maximum transmission centered at 520 nm to a photomultiplier tube (Aminco Cat.…”

Section: Preparation and Characteristics Of Cytochrome Oxidase Liposomesmentioning

confidence: 99%

Comparison of energy-transducing capabilities of the two- and three-subunit cytochromes aa3 from Paracoccus denitrificans and the 13-subunit beef heart enzyme

Hendler

Pardhasaradhi

Reynafarje

et al. 1991

Biophysical Journal

107

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In the accompanying paper, we have shown that the two-subunit cytochrome aa3 isolated from Paracoccus denitrificans displays the same kind of complex and interactive redox behavior as the 13-subunit cytochrome aa3 from beef heart. Therefore, the redox characteristics are not dependent on the additional 11 subunits. In the current work, we have examined the energy-transducing capabilities of both the two- and three-subunit enzymes obtained from Paracoccus denitrificans in relation to that of the 13-unit mammalian enzyme. We have found that in all of the tested functions, which included the development of delta psi and delta pH, and the pumping of protons, that the two-subunit enzyme is at least as efficient as the structurally more complex mammalian enzyme. There is thus a correlation between the complex redox behavior and energy transducing capabilities of the two enzymes. There was also no difference in energy-transducing capabilities between the two- and three-subunit forms of the bacterial enzyme. It seems that only 2 subunits are required for an efficient energy-transducing cytochrome aa3. The most likely role of the additional subunits in the mammalian enzyme, therefore, seems to be in regulation.

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Instrumentation and procedures for real time measurements of proton motive force, membrane potential, ΔpH, proton extrusion, and oxygen uptake in respiring cells and vesicles

Cited by 16 publications

References 14 publications

Direct measurement of the initial proton extrusion to oxygen uptake ratio accompanying succinate oxidation by rat liver mitochondria

Direct measurement of the initial proton extrusion to oxygen uptake ratio accompanying succinate oxidation by rat liver mitochondria

Functional Synergism of the Magainins PGLa and Magainin-2 in Escherichia coli, Tumor Cells and Liposomes

Comparison of energy-transducing capabilities of the two- and three-subunit cytochromes aa3 from Paracoccus denitrificans and the 13-subunit beef heart enzyme

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