Cryo-EM structures reveal ubiquinone-10 binding to mitochondrial complex I and conformational transitions associated with Q-site occupancy

Abstract

Mitochondrial complex I is a central metabolic enzyme that uses the reducing potential of NADH to reduce ubiquinone-10 (Q10) and drive four protons across the inner mitochondrial membrane, powering oxidative phosphorylation. Although many complex I structures are now available, structures of Q10-bound states have remained elusive. Here, we reconstitute mammalian complex I into phospholipid nanodiscs with exogenous Q10. Using cryo-EM, we reveal a Q10 molecule occupying the full length of the Q-binding site in the active (ready-to-go) resting state (plus a matching substrate-free structure) and apply molecular dynamics simulations to propose how the charge states of key residues influence the Q10 binding pose. By comparing ligand-bound and ligand-free forms of the deactive resting state (that require reactivating to catalyse), we begin to define how substrate binding restructures the deactive Q-binding site, providing insights into its physiological and mechanistic relevance.