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Lipid-mediated Protein-protein Interactions Modulate Respiration-driven ATP Synthesis
Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för biokemi och biofysik.ORCID-id: 0000-0003-2930-801X
Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för biokemi och biofysik.
Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för biokemi och biofysik.
Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för biokemi och biofysik.
Vise andre og tillknytning
Rekke forfattare: 62016 (engelsk)Inngår i: Scientific Reports, ISSN 2045-2322, E-ISSN 2045-2322, Vol. 6, artikkel-id 24113Artikkel i tidsskrift (Fagfellevurdert) Published
Abstract [en]

Energy conversion in biological systems is underpinned by membrane-bound proton transporters that generate and maintain a proton electrochemical gradient across the membrane which used, e.g. for generation of ATP by the ATP synthase. Here, we have co-reconstituted the proton pump cytochrome bo3 (ubiquinol oxidase) together with ATP synthase in liposomes and studied the effect of changing the lipid composition on the ATP synthesis activity driven by proton pumping. We found that for 100 nm liposomes, containing 5 of each proteins, the ATP synthesis rates decreased significantly with increasing fractions of DOPA, DOPE, DOPG or cardiolipin added to liposomes made of DOPC; with e.g. 5% DOPG, we observed an almost 50% decrease in the ATP synthesis rate. However, upon increasing the average distance between the proton pumps and ATP synthases, the ATP synthesis rate dropped and the lipid dependence of this activity vanished. The data indicate that protons are transferred along the membrane, between cytochrome bo3 and the ATP synthase, but only at sufficiently high protein densities. We also argue that the local protein density may be modulated by lipid-dependent changes in interactions between the two proteins complexes, which points to a mechanism by which the cell may regulate the overall activity of the respiratory chain.

sted, utgiver, år, opplag, sider
2016. Vol. 6, artikkel-id 24113
HSV kategori
Forskningsprogram
biokemi
Identifikatorer
URN: urn:nbn:se:su:diva-130172DOI: 10.1038/srep24113ISI: 000374219300001PubMedID: 27063297OAI: oai:DiVA.org:su-130172DiVA, id: diva2:927151
Tilgjengelig fra: 2016-05-11 Laget: 2016-05-09 Sist oppdatert: 2020-02-20bibliografisk kontrollert
Inngår i avhandling
1. Modulators of Saccharomyces cerevisiae cytochrome c oxidase: Implications for the regulation of mitochondrial respiration
Åpne denne publikasjonen i ny fane eller vindu >>Modulators of Saccharomyces cerevisiae cytochrome c oxidase: Implications for the regulation of mitochondrial respiration
2018 (engelsk)Doktoravhandling, med artikler (Annet vitenskapelig)
Abstract [en]

Oxidative phosphorylation in mitochondria is performed by enzyme complexes and electron carriers that reside in the inner membrane. It is now generally accepted that these respiratory enzyme complexes assemble into larger so-called supercomplexes. However, it is presently not known why, under which conditions or how these supercomplexes form.

A number of factors of particular importance for the formation of supercomplexes have been identified, such as the Respiratory supercomplex factors (Rcf1 and Rcf2) and cardiolipin. The work presented in this thesis is focused on the characterization of cytochrome c oxidase (CytcO) in mitochondria from Saccharomyces cerevisiae strains in which these components have been removed, with a particular focus on Rcf1. First, we concluded that Rcf1 has an impact on the activity and ligand binding kinetics of CytcO, which upon genetic deletion of rcf1 leads to formation of sub-populations of CytcO with different functionality. Second, we noted that the ability of CytcO to oxidize cytochrome c (cyt. c) depends on the presence of Rcf1. Further, we observed that while CytcO in wild-type mitochondria displayed differences in the oxidation kinetics of cyt. c from horse heart or S. cerevisiae, with the Δrcf1 mitochondria these differences were lost. This observation suggested that Rcf1 interacts with cyt. c. Furthermore, the data showed that in CytcO from Δrcf1 mitochondria heme a3 was altered while heme a was intact.

Using proteo-liposomes of different lipid composition and size we also investigated the influence of lipid head groups on the coupled activity of a quinol oxidase and ATP-synthase. Specifically, we addressed the question if protons are transferred between proton “producers” and “consumers” via lateral proton transfer along the membrane surface or via bulk water. Our data supported the principle of lateral proton transfer.

Lastly, we characterized the ligand binding of yeast flavohemoglobin and concluded that the flavohemoglobin has a population that resides in the intermembrane space of mitochondria, not only in matrix and cytosol as previously suggested.

sted, utgiver, år, opplag, sider
Stockholm: Department of Biochemistry and Biophysics, Stockholm University, 2018. s. 48
Emneord
cytochrome c oxidase, cytochrome c, OXPHOS, membrane protein, kinetics, ligand-binding, electron transfer, Rcf1, respiratory supercomplexes, Saccharomyces cerevisiae
HSV kategori
Forskningsprogram
biokemi
Identifikatorer
urn:nbn:se:su:diva-161515 (URN)978-91-7797-457-4 (ISBN)978-91-7797-456-7 (ISBN)
Disputas
2018-12-13, Magnélisalen, Kemiska övningslaboratoriet, Svante Arrhenius väg 16 B, Stockholm, 10:00 (engelsk)
Opponent
Veileder
Tilgjengelig fra: 2018-11-20 Laget: 2018-10-29 Sist oppdatert: 2018-11-13bibliografisk kontrollert
2. Protein and lipid interactions within the respiratory chain: Studies using membrane-mimetic systems
Åpne denne publikasjonen i ny fane eller vindu >>Protein and lipid interactions within the respiratory chain: Studies using membrane-mimetic systems
2019 (engelsk)Doktoravhandling, med artikler (Annet vitenskapelig)
Abstract [en]

Energy conversion from nutrients to ATP is a vital process in cells. The process, called oxidative phosphorylation (OXPHOS) is performed by a combination of membrane-bound proteins. These proteins have been studied in great detail in the past, however much is still unknown about how they interact with each other. Studying the OXPHOS proteins in their native environment can be difficult due to the complexity of living cells. By isolating parts of the OXPHOS system and inserting them into membrane-mimetic systems it is possible to investigate their functions in a controlled environment.

In the work presented here, we co-reconstituted several of these proteins into liposomes made from synthetic lipids. We demonstrated production of ATP at steady-state conditions with the ATP synthase, driven by proton pumping by cytochrome bo3. Introduction of anionic lipids decreased the coupled activity and we could correlate this effect to weaker interactions between ATP synthase and cytochrome bo3 in the membrane. We also reconstituted cytochrome c oxidase (CytcO) from Saccharomyces cerevisiae with Respiratory supercomplex factor 1 (Rcf1) into liposomes and submitochondrial particles (SMPs). Loss of Rcf1 has previously been found to result in a lower CytcO activity. We found that activity could be restored upon co-reconstitution of CytcO with Rcf1, but only after unfolding and re-folding of the latter, which shows that Rcf1 can adopt two configurations in the membrane.

sted, utgiver, år, opplag, sider
Stockholm: Department of Biochemistry and Biophysics, Stockholm University, 2019. s. 67
Emneord
Cytochrome c oxidase, ATP synthase, Bioenergetics, membrane-mimetics, Rcf1, liposomes, oxidative phosphorylation, lipids, protein-protein interactions
HSV kategori
Forskningsprogram
biokemi
Identifikatorer
urn:nbn:se:su:diva-173617 (URN)978-91-7797-845-9 (ISBN)978-91-7797-846-6 (ISBN)
Disputas
2019-11-22, Magnélisalen, Kemiska övningslaboratoriet, Svante Arrhenius väg 16 B, Stockholm, 10:00 (engelsk)
Opponent
Veileder
Merknad

At the time of the doctoral defense, the following paper was unpublished and had a status as follows: Paper 4: Manuscript.

Tilgjengelig fra: 2019-10-30 Laget: 2019-09-26 Sist oppdatert: 2019-11-27bibliografisk kontrollert

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