Proton Translocations in Isolated Spinach Chloroplasts after Single-turnover Actinic Flashes

1979 ◽  
Vol 6 (3) ◽  
pp. 289
Author(s):  
A.B Hope ◽  
A Morland
Keyword(s):  
Steady State ◽  
Electron Transport ◽  
Electron Transport Chain ◽  
Neutral Red ◽  
Plastoquinone Pool ◽  
Single Turnover ◽  
Transport Chain ◽  
Initial Reduction ◽  
Proton Uptake ◽  
Cresol Red

Proton movements were monitored with cresol red and neutral red. Absorbance changes of these dyes following actinic flashes were measured both for the steady state and for sequences of 6-9 flashes given to separate dark-adapted suspensions of chloroplasts. The cresol red changes corresponded to a steady-state proton uptake (�H*+ss) of 0.0015-0.0017 mol H+/mol Chl per flash (ferricyanide as electron acceptor) or 0.0031 mol H+/mol Chl per flash (methyl viologen). Assuming 600 Chl per electron transport chain, these uptakes corresponded to H+/e- = 1 or 2 respectively. In flash sequences with ferricyanide, a minimum of about 0.6 �H*+ss was noted after flash No. 3. Controls, and considerations of the partitioning of unprotonated neutral red into the thylakoid membrane phase, strongly indicated that neutral red was responding to an acidification of the intrathylakoid aqueous phase. The steady-state signal was interpreted as 2 H+ deposited per electron transport chain per flash (H+/e- = 2), one proton from the oxidation of water and one from the oxidation of plastoquinone. Patterns of proton deposition following sequences of flashes and their change upon adding dibromothymoquinone suggested that (a) protons are produced one at a time in the advance of the 'S-states' (Z*n+ � Z(n+1)+ ); and (b) the precursor to the plastoquinone pool, B, is a special plastoquinone, requiring 1 e- and 1 H+ for its initial reduction but passing on to the plastoquinone pool 2 e- upon the next flash. The proton deposition on flash No. 1 corresponded to a small proportion of reduced B in dark-adapted chloroplasts, as previously postulated.

scholarly journals The function of ubiquinone in Escherichia coli

1970 ◽  
Vol 117 (3) ◽  
pp. 551-562 ◽  
Author(s):  
G. B. Cox ◽  
N. A. Newton ◽  
F. Gibson ◽  
A. M. Snoswell ◽  
J. A. Hamilton
Keyword(s):  
Escherichia Coli ◽  
Electron Spin Resonance ◽  
Steady State ◽  
Electron Transport ◽  
Electron Spin ◽  
Electron Transport Chain ◽  
Electron Spin Resonance Signal ◽  
Resonance Signal ◽  
Transport Chain ◽  
Spin Resonance

1. The function of ubiquinone in Escherichia coli was studied by using whole cells and membrane preparations of normal E. coli and of a mutant lacking ubiquinone. 2. The mutant lacking ubiquinone, strain AN59 (Ubi−), when grown under aerobic conditions, gave an anaerobic type of growth yield and produced large quantities of lactic acid, indicating that ubiquinone plays a vital role in electron transport. 3. NADH and lactate oxidase activities in membranes from strain AN59 (Ubi−) were greatly impaired and activity was restored by the addition of ubiquinone (Q-1). 4. Comparison of the percentage reduction of flavin, cytochrome b1 and cytochrome a2 in the aerobic steady state in membranes from the normal strain (AN62) and strain AN59 (Ubi−) and the effect of respiratory inhibitors on these percentages in membranes from strain AN62 suggest that ubiquinone functions at more than one site in the electron-transport chain. 5. Membranes from strain AN62, in the absence of substrate, showed an electron-spin-resonance signal attributed to ubisemiquinone. The amount of reduced ubiquinone (50%) found after rapid solvent extraction is consistent with the existence of ubiquinone in membranes as a stabilized ubisemiquinone. 6. The effects of piericidin A on membranes from strain AN62 suggest that this inhibitor acts at the ubiquinone sites: thus inhibition of electron transport is reversed by ubiquinone (Q-1); the aerobic steady-state oxidation–reduction levels of flavins and cytochrome b1 in the presence of the inhibitor are raised to values approximating those found in the membranes of strain AN59 (Ubi−); the inhibitor rapidly eliminates the electron-spin-resonance signal attributed to ubisemiquinone and allows slow oxidation of endogenous ubiquinol in the absence of substrate and prevents reduction of ubiquinone in the presence of substrate. It is concluded that piericidin A separates ubiquinone from the remainder of the electron-transport chain. 7. A scheme is proposed in which ubisemiquinone, complexed to an electron carrier, functions in at least two positions in the electron-transport sequence.

scholarly journals Steady-state kinetic analysis of mitochondrial respiratory enzymes from bovine heart mitochondria

2021 ◽  
Vol 64 (1) ◽  
Author(s):  
Dayoung Kim ◽  
Eun Ko ◽  
Moonsung Choi ◽  
Sooim Shin
Keyword(s):  
Steady State ◽  
Electron Transport ◽  
Kinetic Analysis ◽  
Kinetic Parameters ◽  
Complex I ◽  
Electron Transport Chain ◽  
Complex Iii ◽  
Steady State Kinetic ◽  
Transport Chain ◽  
State Kinetic

AbstractMitochondria is a decisive organelle of cells that produces adenosine triphosphate (ATP) by the process of oxidative phosphorylation of the Krebs cycle and the electron transport chain. The electron transport chain system of mitochondria embodies multiple enzyme supercomplexes including complex I to V which located in the inner membrane. Although the simple enzyme activity of some as-isolated complex has been studied so far, the steady-state kinetic analysis of each complex within the form of mitochondrial supercomplex has not been studied in depth. To this end, kinetic parameters of mitochondrial complex I–IV were determined using steady-kinetic analysis using corresponding substrates of them. Catalytic activity and binding affinity between substrates and enzymes were obtained by fitting the data to the Michaelis–Menten equation. Acquired kinetic parameters represented distinctive values depending on the complexes that can be interpreted by the characteristics of the enzymes including the distinction of substrates or the ratio of the enzyme itself under the supercomplex form. The indirect kcat of the mitochondrial enzymes were varied from 0.0609 to 0.334 s−1 in order of complex III, II, I, and IV and Km of substrates were also diverse from 5.1 μM to 12.14 mM. This is the first attempt to get exact kinetic values that should provide profound information to evaluate the mitochondrial function practically in advance.

scholarly journals AB0031 T HELPER 17 CELLS WERE SIGNIFICANTLY DECREASED BY MITOCHONDRIAL ELECTRON TRANSPORT CHAIN COMPLEX INHIBITOR IN PATIENTS WITH RHEUMATOID ARTHRITIS

2020 ◽  
Vol 79 (Suppl 1) ◽  
pp. 1318.2-1318
Author(s):  
H. R. Lee ◽  
S. J. Yoo ◽  
J. Kim ◽  
I. S. Yoo ◽  
C. K. Park ◽  
...  
Keyword(s):  
Rheumatoid Arthritis ◽  
Electron Transport ◽  
Disease Activity ◽  
Th17 Cells ◽  
Electron Transport Chain ◽  
Chain Complex ◽  
Complex Iii ◽  
Mitochondrial Electron Transport Chain ◽  
Transport Chain ◽  
Healthy Control

Background:Reactive oxygen species (ROS) and T helper 17 (TH17) cells have been known to play an important role in the pathogenesis of rheumatoid arthritis (RA). However, the interrelationship between ROS and TH17 remains unclear in RAObjectives:To explore whether ROS affect TH17 cells in peripheral blood mononuclear cells (PBMC) of RA patients, we analyzed ROS expressions among T cell subsets following treatment with mitochondrial electron transport chain complex inhibitors.Methods:Blood samples were collected from 40 RA patients and 10 healthy adult volunteers. RA activity was divided according to clinical parameter DAS28. PBMC cells were obtained from the whole blood using lymphocyte separation medium density gradient centrifugation. Following PBMC was stained with Live/Dead stain dye, cells were incubated with antibodies for CD3, CD4, CD8, and CD25. After fixation and permeabilization, samples were stained with antibodies for FoxP3 and IL-17A. MitoSox were used for mitochondrial specific staining.Results:The frequency of TH17 cells was increased by 4.83 folds in moderate disease activity group (5.1>DAS28≥3.2) of RA patients compared to healthy control. Moderate RA activity patients also showed higher ratio of TH17/Treg than healthy control (3.57 folds). All RA patients had elevated expression of mitochondrial specific ROS than healthy control. When PBMC cells were treated with 2.5uM of antimycin A (mitochondrial electron transport chain complex III inhibitor) for 16 h, the frequency of TH17 cells was significantly decreased.Conclusion:The mitochondrial electron transport chain complex III inhibitor markedly downregulated the frequency of TH17 cells in moderate disease activity patients with RA. These findings provide a novel approach to regulate TH17 function in RA through mitochondrial metabolism related ROS production.References:[1]Szekanecz, Z., et al., New insights in synovial angiogenesis. Joint Bone Spine, 2010. 77(1): p. 13-9.[2]Prevoo, M.L., et al., Modified disease activity scores that include twenty-eight-joint counts. Development and validation in a prospective longitudinal study of patients with rheumatoid arthritis. Arthritis Rheum, 1995. 38(1): p. 44-8.Disclosure of Interests:None declared

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