Repository logo
 

Membrane-domain mutations in respiratory complex I impede catalysis but do not uncouple proton pumping from ubiquinone reduction.

Published version
Peer-reviewed

Repository DOI


Change log

Authors

Jarman, Owen D 

Abstract

Respiratory complex I [NADH:ubiquinone (UQ) oxidoreductase] captures the free energy released from NADH oxidation and UQ reduction to pump four protons across an energy-transducing membrane and power ATP synthesis. Mechanisms for long-range energy coupling in complex I have been proposed from structural data but not yet evaluated by robust biophysical and biochemical analyses. Here, we use the powerful bacterial model system Paracoccus denitrificans to investigate 14 mutations of key residues in the membrane-domain Nqo13/ND4 subunit, defining the rates and reversibility of catalysis and the number of protons pumped per NADH oxidized. We reveal new insights into the roles of highly conserved charged residues in lateral energy transduction, confirm the purely structural role of the Nqo12/ND5 transverse helix, and evaluate a proposed hydrated channel for proton uptake. Importantly, even when catalysis is compromised the enzyme remains strictly coupled (four protons are pumped per NADH oxidized), providing no evidence for escape cycles that circumvent blocked proton-pumping steps.

Description

Acknowledgements: We thank Daniel N. Grba and Justin G. Fedor for useful discussions regarding mutant design, Bozhidar S. Ivanov for help setting up the NanoDSF assay conditions, and John J. Wright for critical reading of the manuscript.

Keywords

NADH:ubiquinone oxidoreductase, biological energy transduction, electron transport chain, proton pumping, respiratory chain

Journal Title

PNAS Nexus

Conference Name

Journal ISSN

2752-6542
2752-6542

Volume Title

1

Publisher

Oxford University Press (OUP)
Sponsorship
MRC (MC_UU_00015/2)
Medical Research Council (MC_UU_00015/7)