| 1. |
- Fedotovskaya, Olga, et al.
(author)
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Identification of a cytochrome bc1-aa3 supercomplex in Rhodobacter sphaeroides
- 2021
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In: Biochimica et Biophysica Acta - Bioenergetics. - : Elsevier BV. - 0005-2728 .- 1879-2650. ; 1862:8
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Journal article (peer-reviewed)abstract
- Respiration is carried out by a series of membrane-bound complexes in the inner mitochondrial membrane or in the cytoplasmic membrane of bacteria. Increasing evidence shows that these complexes organize into larger supercomplexes. In this work, we identified a supercomplex composed of cytochrome (cyt.) bc1 and aa3-type cyt. c oxidase in Rhodobacter sphaeroides. We purified the supercomplex using a His-tag on either of these complexes. The results from activity assays, native and denaturing PAGE, size exclusion chromatography, electron microscopy, optical absorption spectroscopy and kinetic studies on the purified samples support the formation and coupled quinol oxidation:O2 reduction activity of the cyt. bc1-aa3 supercomplex. The potential role of the membrane-anchored cyt. cy as a component in supercomplexes was also investigated.
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| 2. |
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| 3. |
- Graf, Simone, et al.
(author)
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Rapid Electron Transfer within the III-IV Supercomplex in Corynebacterium glutamicum
- 2016
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In: Scientific Reports. - : Springer Science and Business Media LLC. - 2045-2322. ; 6
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Journal article (peer-reviewed)abstract
- Complex III in C. glutamicum has an unusual di-heme cyt.c(1) and it co-purifies with complex IV in a supercomplex. Here, we investigated the kinetics of electron transfer within this supercomplex and in the cyt.aa(3) alone (cyt.bc(1) was removed genetically). In the reaction of the reduced cyt.aa(3) with O-2, we identified the same sequence of events as with other A-type oxidases. However, even though this reaction is associated with proton uptake, no pH dependence was observed in the kinetics. For the cyt. bc(1)-cyt.aa(3) supercomplex, we observed that electrons from the c-hemes were transferred to CuA with time constants 0.1-1 ms. The b-hemes were oxidized with a time constant of 6.5 ms, indicating that this electron transfer is rate-limiting for the overall quinol oxidation/O-2 reduction activity (similar to 210 e(-)/s). Furthermore, electron transfer from externally added cyt.c to cyt.aa(3) was significantly faster upon removal of cyt.bc(1) from the supercomplex, suggesting that one of the c-hemes occupies a position near Cu-A. In conclusion, isolation of the III-IV-supercomplex allowed us to investigate the kinetics of electron transfer from the b-hemes, via the di-heme cyt.c(1) and heme a to the heme a(3)-Cu-B catalytic site of cyt.aa(3).
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| 4. |
- Król, Sylwia, et al.
(author)
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Electron and proton transfer in the M. smegmatis III2IV2 supercomplex
- 2022
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In: Biochimica et Biophysica Acta - Bioenergetics. - : Elsevier BV. - 0005-2728 .- 1879-2650. ; 1863:7
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Journal article (peer-reviewed)abstract
- The M. smegmatis respiratory III2IV2 supercomplex consists of a complex III (CIII) dimer flanked on each side by a complex IV (CIV) monomer, electronically connected by a di-heme cyt. cc subunit of CIII. The supercomplex displays a quinol oxidation‑oxygen reduction activity of ~90 e−/s. In the current work we have investigated the kinetics of electron and proton transfer upon reaction of the reduced supercomplex with molecular oxygen. The data show that, as with canonical CIV, oxidation of reduced CIV at pH 7 occurs in three resolved components with time constants ~30 μs, 100 μs and 4 ms, associated with the formation of the so-called peroxy (P), ferryl (F) and oxidized (O) intermediates, respectively. Electron transfer from cyt. cc to the primary electron acceptor of CIV, CuA, displays a time constant of ≤100 μs, while re-reduction of cyt. cc by heme b occurs with a time constant of ~4 ms. In contrast to canonical CIV, neither the P → F nor the F → O reactions are pH dependent, but the P → F reaction displays a H/D kinetic isotope effect of ~3. Proton uptake through the D pathway in CIV displays a single time constant of ~4 ms, i.e. a factor of ~40 slower than with canonical CIV. The slowed proton uptake kinetics and absence of pH dependence are attributed to binding of a loop from the QcrB subunit of CIII at the D proton pathway of CIV. Hence, the data suggest that function of CIV is modulated by way of supramolecular interactions with CIII.
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| 5. |
- Toth, Alexandra, 1989-, et al.
(author)
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Membrane tethering of cytochrome c accelerates apoptotic cell death in yeast
- 2020
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In: Cell Death and Disease. - : Springer Science and Business Media LLC. - 2041-4889. ; 11:9
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Journal article (peer-reviewed)abstract
- Intrinsic apoptosis as a modality of regulated cell death is intimately linked to permeabilization of the outer mitochondrial membrane and subsequent release of the protein cytochrome c into the cytosol, where it can participate in caspase activation via apoptosome formation. Interestingly, cytochrome c release is an ancient feature of regulated cell death even in unicellular eukaryotes that do not contain an apoptosome. Therefore, it was speculated that cytochrome c release might have an additional, more fundamental role for cell death signalling, because its absence from mitochondria disrupts oxidative phosphorylation. Here, we permanently anchored cytochrome c with a transmembrane segment to the inner mitochondrial membrane of the yeast Saccharomyces cerevisiae, thereby inhibiting its release from mitochondria during regulated cell death. This cytochrome c retains respiratory growth and correct assembly of mitochondrial respiratory chain supercomplexes. However, membrane anchoring leads to a sensitisation to acetic acid-induced cell death and increased oxidative stress, a compensatory elevation of cellular oxygen-consumption in aged cells and a decreased chronological lifespan. We therefore conclude that loss of cytochrome c from mitochondria during regulated cell death and the subsequent disruption of oxidative phosphorylation is not required for efficient execution of cell death in yeast, and that mobility of cytochrome c within the mitochondrial intermembrane space confers a fitness advantage that overcomes a potential role in regulated cell death signalling in the absence of an apoptosome.
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| 6. |
- Toth, Alexandra, et al.
(author)
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Membrane-tethering of cytochrome c accelerates regulated cell death in yeast
- 2020
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In: Cell Death and Disease. - : Springer Nature. - 2041-4889. ; 11:9
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Journal article (peer-reviewed)abstract
- Intrinsic apoptosis as a modality of regulated cell death is intimately linked to permeabilization of the outer mitochondrial membrane and subsequent release of the protein cytochrome c into the cytosol, where it can participate in caspase activation via apoptosome formation. Interestingly, cytochrome c release is an ancient feature of regulated cell death even in unicellular eukaryotes that do not contain an apoptosome. Therefore, it was speculated that cytochrome c release might have an additional, more fundamental role for cell death signalling, because its absence from mitochondria disrupts oxidative phosphorylation. Here, we permanently anchored cytochrome c with a transmembrane segment to the inner mitochondrial membrane of the yeast Saccharomyces cerevisiae, thereby inhibiting its release from mitochondria during regulated cell death. This cytochrome c retains respiratory growth and correct assembly of mitochondrial respiratory chain supercomplexes. However, membrane anchoring leads to a sensitisation to acetic acid-induced cell death and increased oxidative stress, a compensatory elevation of cellular oxygen-consumption in aged cells and a decreased chronological lifespan. We therefore conclude that loss of cytochrome c from mitochondria during regulated cell death and the subsequent disruption of oxidative phosphorylation is not required for efficient execution of cell death in yeast, and that mobility of cytochrome c within the mitochondrial intermembrane space confers a fitness advantage that overcomes a potential role in regulated cell death signalling in the absence of an apoptosome.
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| 7. |
- Wiseman, Benjamin, et al.
(author)
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Structure of a functional obligate complex III2IV2 respiratory supercomplex from Mycobacterium smegmatis
- 2018
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In: Nature Structural & Molecular Biology. - : Springer Science and Business Media LLC. - 1545-9993 .- 1545-9985. ; 25:12, s. 1128-1136
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Journal article (peer-reviewed)abstract
- In the mycobacterial electron-transport chain, respiratory complex III passes electrons from menaquinol to complex IV, which in turn reduces oxygen, the terminal acceptor. Electron transfer is coupled to transmembrane proton translocation, thus establishing the electrochemical proton gradient that drives ATP synthesis. We isolated, biochemically characterized, and determined the structure of the obligate III2IV2 supercomplex from Mycobacterium smegmatis, a model for Mycobacterium tuberculosis. The supercomplex has quinol:O-2 oxidoreductase activity without exogenous cytochrome c and includes a superoxide dismutase subunit that may detoxify reactive oxygen species produced during respiration. We found menaquinone bound in both the Q(o) and Q(i) sites of complex III. The complex III-intrinsic diheme cytochrome cc subunit, which functionally replaces both cytochrome c(1) and soluble cytochrome c in canonical electron-transport chains, displays two conformations: one in which it provides a direct electronic link to complex IV and another in which it serves as an electrical switch interrupting the connection.
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| 8. |
- Yamada, Takashi, et al.
(author)
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Nitrosative modifications of the Ca2+ release complex and actin underlie arthritis-induced muscle weakness.
- 2015
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In: Annals of the Rheumatic Diseases. - : BMJ. - 1468-2060 .- 0003-4967. ; 74:10, s. 1907-1914
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Journal article (peer-reviewed)abstract
- Skeletal muscle weakness is a prominent clinical feature in patients with rheumatoid arthritis (RA), but the underlying mechanism(s) is unknown. Here we investigate the mechanisms behind arthritis-induced skeletal muscle weakness with special focus on the role of nitrosative stress on intracellular Ca(2+) handling and specific force production.
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