Change search
ReferencesLink to record
Permanent link

Direct link
Mimicking respiratory phosphorylation using purified enzymes
Stockholm University, Faculty of Science, Department of Biochemistry and Biophysics.
Stockholm University, Faculty of Science, Department of Biochemistry and Biophysics.
Number of Authors: 4
2016 (English)In: Biochimica et Biophysica Acta - Bioenergetics, ISSN 0005-2728, E-ISSN 1879-2650, Vol. 1857, no 4, 321-331 p.Article in journal (Refereed) Published
Abstract [en]

The enzymes of oxidative phosphorylation is a striking example of the functional association of multiple enzyme complexes, working together to form ATP from cellular reducing equivalents. These complexes, such as cytochrome c oxidase or the ATP synthase, are typically investigated individually and therefore, their functional interplay is not well understood. Here, we present methodology that allows the co-reconstitution of purified terminal oxidases and ATP synthases in synthetic liposomes. The enzymes are functionally coupled via proton translocation where upon addition of reducing equivalents the oxidase creates and maintains a transmembrane electrochemical proton gradient that energizes the synthesis of ATP by the F1F0 ATP synthase. The method has been tested with the ATP synthases from Escherichia coli and spinach chloroplasts, and with the quinol and cytochrome c oxidases from E. coli and Rhodobacter sphaeroides, respectively. Unlike in experiments with the ATP synthase reconstituted alone, the setup allows in vitro ATP synthesis under steady state conditions, with rates up to 90 ATP x s(-1) x enzyme(-1). We have also used the novel system to study the phenomenon of mild uncoupling as observed in mitochondria upon addition of low concentrations of ionophores (e.g. FCCP, SF6847) and the recoupling effect of 6-ketocholestanol. While we could reproduce the described effects, our data with the in vitro system does not support the idea of a direct interaction between a mitochondrial protein and the uncoupling agents as proposed earlier.

Place, publisher, year, edition, pages
2016. Vol. 1857, no 4, 321-331 p.
Keyword [en]
ATP synthesis, Respiratory chain, Liposomes, Mild uncoupling, Ionophore, Lateral proton diffusion
National Category
Biological Sciences
Identifiers
URN: urn:nbn:se:su:diva-129914DOI: 10.1016/j.bbabio.2015.12.007ISI: 000372675600001PubMedID: 26707617OAI: oai:DiVA.org:su-129914DiVA: diva2:925778
Available from: 2016-05-03 Created: 2016-05-03 Last updated: 2016-05-03Bibliographically approved

Open Access in DiVA

No full text

Other links

Publisher's full textPubMed

Search in DiVA

By author/editor
Nilsson, TobiasBrzezinski, Peter
By organisation
Department of Biochemistry and Biophysics
In the same journal
Biochimica et Biophysica Acta - Bioenergetics
Biological Sciences

Search outside of DiVA

GoogleGoogle Scholar
The number of downloads is the sum of all downloads of full texts. It may include eg previous versions that are now no longer available

Altmetric score

Total: 29 hits
ReferencesLink to record
Permanent link

Direct link