Issue 6, 2023

Using light scattering to assess how phospholipid–protein interactions affect complex I functionality in liposomes

Abstract

Complex I is an essential membrane protein in respiration, oxidising NADH and reducing ubiquinone to contribute to the proton-motive force that powers ATP synthesis. Liposomes provide an attractive platform to investigate complex I in a phospholipid membrane with the native hydrophobic ubiquinone substrate and proton transport across the membrane, but without convoluting contributions from other proteins present in the native mitochondrial inner membrane. Here, we use dynamic and electrophoretic light scattering techniques (DLS and ELS) to show how physical parameters, in particular the zeta potential (ζ-potential), correlate strongly with the biochemical functionality of complex I-containing proteoliposomes. We find that cardiolipin plays a crucial role in the reconstitution and functioning of complex I and that, as a highly charged lipid, it acts as a sensitive reporter on the biochemical competence of proteoliposomes in ELS measurements. We show that the change in ζ-potential between liposomes and proteoliposomes correlates linearly with protein retention and catalytic oxidoreduction activity of complex I. These correlations are dependent on the presence of cardiolipin, but are otherwise independent of the liposome lipid composition. Moreover, changes in the ζ-potential are sensitive to the proton motive force established upon proton pumping by complex I, thereby constituting a complementary technique to established biochemical assays. ELS measurements may thus serve as a more widely useful tool to investigate membrane proteins in lipid systems, especially those that contain charged lipids.

Graphical abstract: Using light scattering to assess how phospholipid–protein interactions affect complex I functionality in liposomes

Supplementary files

Article information

Article type
Paper
Submitted
29 6月 2022
Accepted
20 3月 2023
First published
20 3月 2023
This article is Open Access
Creative Commons BY license

RSC Chem. Biol., 2023,4, 386-398

Using light scattering to assess how phospholipid–protein interactions affect complex I functionality in liposomes

J. Eisermann, J. J. Wright, J. D. E. T. Wilton-Ely, J. Hirst and M. M. Roessler, RSC Chem. Biol., 2023, 4, 386 DOI: 10.1039/D2CB00158F

This article is licensed under a Creative Commons Attribution 3.0 Unported Licence. You can use material from this article in other publications without requesting further permissions from the RSC, provided that the correct acknowledgement is given.

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