Complex I (NADH:ubiquinone oxidoreductase) uses the reducing potential of NADH to drive protons across the energy-transducing inner membrane and power oxidative phosphorylation in mammalian mitochondria. Recently, cryoEM analyses have produced close-to-complete models of all 45 subunits in the bovine, ovine and porcine complexes, and identifed two states relevant to complex I in ischemia-reperfusion injury. Here, we describe the 3.3-Å structure of complex I from mouse heart mitochondria, a biomedically-relevant model system, in the ‘active’ state. We reveal a nucleotide bound in subunit NDUFA10, a nucleoside-kinase homolog, and define mechanistically-critical elements in the mammalian enzyme. By comparisons with a 3.9-Å structure of the ‘deactive’ state and with known bacterial structures we identify differences in helical geometry in the membrane domain that occur upon activation, or that alter the positions of catalytically-important charged residues. Our results demonstrate the capability of cryoEM analyses to challenge and develop mechanistic models for mammalian complex I.

Bacterial efflux pumps confer multidrug resistance by transporting diverse antibiotics from the cell. In Gram-negative bacteria, some of these pumps form multi-protein assemblies that span the cell envelope. Here, we report the near-atomic resolution cryoEM structures of the Escherichia coli AcrAB-TolC multidrug efflux pump in resting and drug transport states, revealing a quaternary structural switch that allosterically couples and synchronizes initial ligand binding with channel opening. Within the transport-activated state, the channel remains open even though the pump cycles through three distinct conformations. Collectively, our data provide a dynamic mechanism for the assembly and operation of the AcrAB-TolC pump.DOI: http://dx.doi.org/10.7554/eLife.24905.001

Respiratory chain dysfunction plays an important role in human disease and aging. It is now well established that the individual respiratory complexes can be organized into supercomplexes, and structures for these macromolecular assemblies, determined by electron cryo-microscopy, have been described recently. Nevertheless, the reason why supercomplexes exist remains an enigma. The widely held view that they enhance catalysis by channeling substrates is challenged by both structural and biophysical information. Here, we evaluate and discuss data and hypotheses on the structures, roles, and assembly of respiratory-chain supercomplexes and propose a future research agenda to address unanswered questions.