TY - JOUR
T1 - Identification of a cytochrome bc1-aa3 supercomplex in Rhodobacter sphaeroides
AU - Fedotovskaya, Olga
AU - Albertsson, Ingrid
AU - Nordlund, Gustav
AU - Hong, Sangjin
AU - Gennis, Robert B.
AU - Brzezinski, Peter
AU - Ädelroth, Pia
N1 - Funding Information:
This work was supported by grants from the Knut and Alice Wallenberg Foundation (grants number 2013.0006 and 2019.0043) and the Swedish Research Council to PB (VR grant 2018-04619) and PÄ (VR grant number 2019-04124). We are grateful to Dan Daley (Stockholm University) for help with BN-PAGE, to Dan Sjöstrand (Stockholm University) for help with the Yarra 4000 SEC column, and to Agnes Moe and Ben Wiseman (Stockholm University) for help with the negative stain image processing. The mass spectrometry analysis was performed by the mass spectrometry proteomics facility at Uppsala University.
Funding Information:
Upon addition of quinol to SCbc 1 and SCaa 3 we observed coupled QH 2 oxidation-O 2 reduction, which indicated an uninterrupted electron flow from the QH 2 through the cyt. bc 1 to the aa 3 -CytcO. Since this activity was observed in the absence of added cyt. c, there must be an alternative link between the complexes, and we suggest this link to be the MA-cyt. c y . The presence of MA-cyt. c y was supported by the excess heme c in optical spectra ( Fig. 1 ), by the additional heme-stained band of ~20 kDa ( Fig. 5 ), and by mass spectrometry (Table S5). We found no support for the presence of the soluble cyt. c 2 in the samples.
Publisher Copyright:
© 2021 The Authors
PY - 2021/8/1
Y1 - 2021/8/1
N2 - 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.
AB - 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.
KW - Bioenergetics
KW - Cytochrome bc complex
KW - Electron transfer
KW - Respiratory supercomplex
KW - aa-type cytochrome c oxidase
KW - flow-flash
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U2 - 10.1016/j.bbabio.2021.148433
DO - 10.1016/j.bbabio.2021.148433
M3 - Article
C2 - 33932366
AN - SCOPUS:85105322483
SN - 0005-2728
VL - 1862
JO - Biochimica et Biophysica Acta - Bioenergetics
JF - Biochimica et Biophysica Acta - Bioenergetics
IS - 8
M1 - 148433
ER -