@article{1226, abstract = {Mitochondrial complex I (also known as NADH:ubiquinone oxidoreductase) contributes to cellular energy production by transferring electrons from NADH to ubiquinone coupled to proton translocation across the membrane. It is the largest protein assembly of the respiratory chain with a total mass of 970 kilodaltons. Here we present a nearly complete atomic structure of ovine (Ovis aries) mitochondrial complex I at 3.9 Å resolution, solved by cryo-electron microscopy with cross-linking and mass-spectrometry mapping experiments. All 14 conserved core subunits and 31 mitochondria-specific supernumerary subunits are resolved within the L-shaped molecule. The hydrophilic matrix arm comprises flavin mononucleotide and 8 iron-sulfur clusters involved in electron transfer, and the membrane arm contains 78 transmembrane helices, mostly contributed by antiporter-like subunits involved in proton translocation. Supernumerary subunits form an interlinked, stabilizing shell around the conserved core. Tightly bound lipids (including cardiolipins) further stabilize interactions between the hydrophobic subunits. Subunits with possible regulatory roles contain additional cofactors, NADPH and two phosphopantetheine molecules, which are shown to be involved in inter-subunit interactions. We observe two different conformations of the complex, which may be related to the conformationally driven coupling mechanism and to the active-deactive transition of the enzyme. Our structure provides insight into the mechanism, assembly, maturation and dysfunction of mitochondrial complex I, and allows detailed molecular analysis of disease-causing mutations.}, author = {Fiedorczuk, Karol and Letts, James A and Degliesposti, Gianluca and Kaszuba, Karol and Skehel, Mark and Sazanov, Leonid A}, journal = {Nature}, number = {7625}, pages = {406 -- 410}, publisher = {Nature Publishing Group}, title = {{Atomic structure of the entire mammalian mitochondrial complex i}}, doi = {10.1038/nature19794}, volume = {538}, year = {2016}, } @article{1232, abstract = {Mitochondrial electron transport chain complexes are organized into supercomplexes responsible for carrying out cellular respiration. Here we present three architectures of mammalian (ovine) supercomplexes determined by cryo-electron microscopy. We identify two distinct arrangements of supercomplex CICIII 2 CIV (the respirasome) - a major 'tight' form and a minor 'loose' form (resolved at the resolution of 5.8 Å and 6.7 Å, respectively), which may represent different stages in supercomplex assembly or disassembly. We have also determined an architecture of supercomplex CICIII 2 at 7.8 Å resolution. All observed density can be attributed to the known 80 subunits of the individual complexes, including 132 transmembrane helices. The individual complexes form tight interactions that vary between the architectures, with complex IV subunit COX7a switching contact from complex III to complex I. The arrangement of active sites within the supercomplex may help control reactive oxygen species production. To our knowledge, these are the first complete architectures of the dominant, physiologically relevant state of the electron transport chain.}, author = {Letts, James A and Fiedorczuk, Karol and Sazanov, Leonid A}, journal = {Nature}, number = {7622}, pages = {644 -- 648}, publisher = {Nature Publishing Group}, title = {{The architecture of respiratory supercomplexes}}, doi = {10.1038/nature19774}, volume = {537}, year = {2016}, } @article{1276, abstract = {The cytochrome (cyt) bc 1 complex is an integral component of the respiratory electron transfer chain sustaining the energy needs of organisms ranging from humans to bacteria. Due to its ubiquitous role in the energy metabolism, both the oxidation and reduction of the enzyme's substrate co-enzyme Q has been studied vigorously. Here, this vast amount of data is reassessed after probing the substrate reduction steps at the Q i-site of the cyt bc 1 complex of Rhodobacter capsulatus using atomistic molecular dynamics simulations. The simulations suggest that the Lys251 side chain could rotate into the Q i-site to facilitate binding of half-protonated semiquinone-a reaction intermediate that is potentially formed during substrate reduction. At this bent pose, the Lys251 forms a salt bridge with the Asp252, thus making direct proton transfer possible. In the neutral state, the lysine side chain stays close to the conserved binding location of cardiolipin (CL). This back-and-forth motion between the CL and Asp252 indicates that Lys251 functions as a proton shuttle controlled by pH-dependent negative feedback. The CL/K/D switching, which represents a refinement to the previously described CL/K pathway, fine-tunes the proton transfer process. Lastly, the simulation data was used to formulate a mechanism for reducing the substrate at the Q i-site.}, author = {Postila, Pekka and Kaszuba, Karol and Kuleta, Patryk and Vattulainen, Ilpo and Sarewicz, Marcin and Osyczka, Artur and Róg, Tomasz}, journal = {Scientific Reports}, publisher = {Nature Publishing Group}, title = {{Atomistic determinants of co-enzyme Q reduction at the Qi-site of the cytochrome bc1 complex}}, doi = {10.1038/srep33607}, volume = {6}, year = {2016}, } @article{1288, abstract = {Respiratory complex I transfers electrons from NADH to quinone, utilizing the reaction energy to translocate protons across the membrane. It is a key enzyme of the respiratory chain of many prokaryotic and most eukaryotic organisms. The reversible NADH oxidation reaction is facilitated in complex I by non-covalently bound flavin mononucleotide (FMN). Here we report that the catalytic activity of E. coli complex I with artificial electron acceptors potassium ferricyanide (FeCy) and hexaamineruthenium (HAR) is significantly inhibited in the enzyme pre-reduced by NADH. Further, we demonstrate that the inhibition is caused by reversible dissociation of FMN. The binding constant (Kd) for FMN increases from the femto- or picomolar range in oxidized complex I to the nanomolar range in the NADH reduced enzyme, with an FMN dissociation time constant of ~ 5 s. The oxidation state of complex I, rather than that of FMN, proved critical to the dissociation. Such dissociation is not observed with the T. thermophilus enzyme and our analysis suggests that the difference may be due to the unusually high redox potential of Fe-S cluster N1a in E. coli. It is possible that the enzyme attenuates ROS production in vivo by releasing FMN under highly reducing conditions.}, author = {Holt, Peter and Efremov, Rouslan and Nakamaru Ogiso, Eiko and Sazanov, Leonid A}, journal = {Biochimica et Biophysica Acta - Bioenergetics}, number = {11}, pages = {1777 -- 1785}, publisher = {Elsevier}, title = {{Reversible FMN dissociation from Escherichia coli respiratory complex I}}, doi = {10.1016/j.bbabio.2016.08.008}, volume = {1857}, year = {2016}, } @article{1638, abstract = {The mitochondrial respiratory chain, also known as the electron transport chain (ETC), is crucial to life, and energy production in the form of ATP is the main mitochondrial function. Three proton-translocating enzymes of the ETC, namely complexes I, III and IV, generate proton motive force, which in turn drives ATP synthase (complex V). The atomic structures and basic mechanisms of most respiratory complexes have previously been established, with the exception of complex I, the largest complex in the ETC. Recently, the crystal structure of the entire complex I was solved using a bacterial enzyme. The structure provided novel insights into the core architecture of the complex, the electron transfer and proton translocation pathways, as well as the mechanism that couples these two processes.}, author = {Sazanov, Leonid A}, journal = {Nature Reviews Molecular Cell Biology}, number = {6}, pages = {375 -- 388}, publisher = {Nature Publishing Group}, title = {{A giant molecular proton pump: structure and mechanism of respiratory complex I}}, doi = {10.1038/nrm3997}, volume = {16}, year = {2015}, } @article{1683, abstract = {The 1 MDa, 45-subunit proton-pumping NADH-ubiquinone oxidoreductase (complex I) is the largest complex of the mitochondrial electron transport chain. The molecular mechanism of complex I is central to the metabolism of cells, but has yet to be fully characterized. The last two years have seen steady progress towards this goal with the first atomic-resolution structure of the entire bacterial complex I, a 5 Å cryo-electron microscopy map of bovine mitochondrial complex I and a ∼3.8 Å resolution X-ray crystallographic study of mitochondrial complex I from yeast Yarrowia lipotytica. In this review we will discuss what we have learned from these studies and what remains to be elucidated.}, author = {Letts, Jame A and Sazanov, Leonid A}, journal = {Current Opinion in Structural Biology}, number = {8}, pages = {135 -- 145}, publisher = {Elsevier}, title = {{Gaining mass: The structure of respiratory complex I-from bacterial towards mitochondrial versions}}, doi = {10.1016/j.sbi.2015.08.008}, volume = {33}, year = {2015}, }