Cryo-EM structures of complex I from mouse heart mitochondria in two biochemically defined states

Agip, Ahmed-Noor A.; Blaza, James N.; Bridges, Hannah R.; Viscomi, Carlo; Rawson, Shaun; Muench, Stephen P.; Hirst, Judy

doi:10.1038/s41594-018-0073-1

Cited by 214 publications

(487 citation statements)

References 72 publications

(170 reference statements)

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“…In plant, 25 proteins shared with aerobic yeast or mammals are present. Similarly to the previously obtained complex I structure 4,6,9,16 , the supernumerary subunits, specific to mitochondrial complex I, form a shell around the core subunits, adding nearly 400kDa of proteins to the conserved subunits (Extended Data Fig. 6).…”

Section: General Descriptionsupporting

confidence: 61%

Specific features and assembly of the plant mitochondrial complex I revealed by cryo-EM

Soufari

Parrot

Kuhn

et al. 2020

Preprint

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Mitochondria are the powerhouses of eukaryotic cells and the site of essential metabolic reactions. Their main purpose is to maintain the high ATP/ADP ratio that is required to fuel the countless biochemical reactions taking place in eukaryotic cells 1 . This high ATP/ADP ratio is maintained through oxidative phosphorylation (OXPHOS). Complex I or NADH:ubiquinone oxidoreductase is the main entry site for electrons into the mitochondrial respiratory chain and constitutes the largest of the respiratory complexes 2 . Its structure and composition varies across eukaryotes species. However, high resolution structures are available only for one group of eukaryotes, opisthokonts 3-6 . In plants, only biochemical studies were carried out, already hinting the peculiar composition of complex I in the green lineage. Here, we report several cryoelectron microscopy structures of the plant mitochondrial complex I at near-atomic resolution. We describe the structure and composition of the plant complex I including the plant-specific additional domain composed by carbonic anhydrase proteins. We show that the carbonic anhydrase is an heterotrimeric complex with only one conserved active site. This domain is crucial for the overall stability of complex I as well as a peculiar lipid complex composed cardiolipin and phosphatidylinositols. Moreover we also describe the structure of one of the plant-specific complex I assembly intermediate, lacking the whole PD module, in presence of the maturation factor GLDH. GLDH prevents the binding of the plant specific P1 protein, responsible for the linkage of the PP to the PD module. Finally, as the carbonic anhydrase domain is likely to be associated with complex I from numerous other known eukaryotes, we propose that our structure unveils an ancestral-like organization of mitochondrial complex I.Complex I is the largest multimeric enzyme of the respiratory chain, composed of more than 40 protein subunits. 14 are strictly conserved proteins subunits, vestige of complex I of bacterial origin 7 . The additional subunits, referred to as supernumerary subunits, were acquired during eukaryotes evolution. Part of these additional subunits are conserved among other eukaryotes and play essential roles for the structure, function and the association with the other respiratory chain complexes, eg. to assemble into respirasome in animals 8 . Recently, several high resolution 3D structures of the complete mitochondrial complex I of opisthokonts were determined by cryo-EM in mammalian species 4-6 and in the aerobic yeast Yarrowia lipolytica 3,9 , revealing the organization of their additional specie-specific subunits. However, in plants, even though extensive biochemical characterization was conducted 10,11 , high-resolution structures of mitochondrial complex I are yet to be derived. Early negative staining studies 12 revealed the presence of a large additional membrane attached domain, absent from animal and yeast species, hinting at the peculiar structure and composition of the plant complex I.In order...

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Section: General Descriptionsupporting

confidence: 61%

Specific features and assembly of the plant mitochondrial complex I revealed by cryo-EM

Soufari

Parrot

Kuhn

et al. 2020

Preprint

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“…The Q-chamber geometry and the free volume available to accommodate Q inside are similar among all determined active structures. [8][9][10][11]13 Although complex I has been cocrystallized with small Q analogs, 8,9 it is still unclear how natural substrates with long isoprenoid chains such as ubiquinone-10 in cyan and Nqo8 in pale green that form the Q-chamber with Q 10 colored in green and red alternating for each isoprenoid unit. Elements of secondary structure and residues (green sticks and pink spheres) discussed in the text are indicated.…”

Section: Introductionmentioning

confidence: 99%

Balanced internal hydration discriminates substrate binding to respiratory complex I

Teixeira

Arantes

2019

Preprint

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Molecular recognition of the amphiphilic electron carrier ubiquinone (Q) by respiratory complexes is a fundamental part of electron transfer chains in mitochondria and many bacteria. The primary respiratory complex I binds Q in a long and narrow protein chamber to catalyse its reduction. But, the binding mechanism and the role of chamber hydration in substrate selectivity and stability are unclear. Here, large-scale atomistic molecular dynamics simulations and estimated free energy profiles are used to characterize in detail the binding mechanism to complex I of Q with short and with long isoprenoid tails. A highly stable binding site with two different poses near the chamber exit and a secondary reactive site near the N2 iron-sulfur cluster are found which may lead to an alternative Q redox chemistry and help to explain complex I reactivity. The binding energetics depends mainly on polar interactions of the Q-head and on the counterbalanced hydration of Q-tail isoprenoid units and hydrophobic residues inside the protein chamber. Selectivity upon variation of tail length arises by shifting the hydration balance. This internal hydration mechanism may have implications for binding of amphiphilic molecules to cavities in other membrane proteins.

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“…The X-ray crystallographic structures of complex I from Thermus thermophilus (5) and Yarrowia lipolytica (6) were modeled at resolutions of 3.3 and 3.6 Å, respectively. The entire structures of mammalian complex I, including all 45 subunits (31 of which are the supernumerary subunits), from bovine (Bos taurus) (7,8), ovine (Ovis aries) (9), porcine (Sus scrofa domesticus) (10), and mouse (Mus musculus) (11) hearts and human (Homo sapiens) HEK293F cells (12) were modeled by single-particle cryoelectron microscopy (cryo-EM). One of the striking findings obtained with T. thermophilus complex I (5) is the identification of a long and narrow channel, which extends from the membrane interior to the Fe-S cluster N2 (ϳ30 Å long) and is a completely enclosed tunnel with only a narrow entry point (ϳ3 ϫ 5 Å diameter) for quinone/inhibitors; however, this has not yet been confirmed experimentally.…”

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confidence: 99%

“…From this, the yeast and ovine enzymes were supposed to be in the deactive state. Hirst and co-workers (8,11) recently reported that the structural changes accompanying deactivation may be common to the bovine and mouse enzymes. Considering the unusually long substrate-binding channel, definitions of how UQs of varying isoprenyl chain length (UQ 1 -UQ 10 ) enter and transit the channel to be reduced, thereby eliciting the same proton-pumping stoichiometry, remain elusive (13,14).…”

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confidence: 99%

Exploring the quinone/inhibitor-binding pocket in mitochondrial respiratory complex I by chemical biology approaches

Uno¹,

Kimura²,

Murai

et al. 2019

Journal of Biological Chemistry

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Edited by Ruma Banerjee NADH-quinone oxidoreductase (respiratory complex I) couples NADH-to-quinone electron transfer to the translocation of protons across the membrane. Even though the architecture of the quinone-access channel in the enzyme has been modeled by X-ray crystallography and cryo-EM, conflicting findings raise the question whether the models fully reflect physiologically relevant states present throughout the catalytic cycle. To gain further insights into the structural features of the binding pocket for quinone/inhibitor, we performed chemical biology experiments using bovine heart sub-mitochondrial particles. We synthesized ubiquinones that are oversized (SF-UQs) or lipid-like (PC-UQs) and are highly unlikely to enter and transit the predicted narrow channel. We found that SF-UQs and PC-UQs can be catalytically reduced by complex I, albeit only at moderate or low rates. Moreover, quinone-site inhibitors completely blocked the catalytic reduction and the membrane potential formation coupled to this reduction. Photoaffinity-labeling experiments revealed that amiloride-type inhibitors bind to the interfacial domain of multiple core subunits (49 kDa, ND1, and PSST) and the 39-kDa supernumerary subunit, although the latter does not make up the channel cavity in the current models. The binding of amilorides to the multiple target subunits was remarkably suppressed by other quinone-site inhibitors and SF-UQs. Taken together, the present results are difficult to reconcile with the current channel models. On the basis of comprehensive interpretations of the present results and of previous findings, we discuss the physiological relevance of these models. Figure 2. Structures of photoreactive amilorides synthesized in this study. PRA1 and PRA2 used in our previous work (17) are also shown. A photolabile azido group attached to each compound is indicated by a gray circle. The concentration in parentheses is the average IC 50 value, which is the molar concentration (nanomolar) needed to reduce the control NADH oxidase activity in bovine heart SMPs (30 g of proteins/ml) by 50%. As a reference, the IC 50 value of bullatacin was 1.1 (Ϯ 0.09) nM under the same experimental conditions. Quinone/inhibitor-binding pocket in respiratory complex I

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Cryo-EM structures of complex I from mouse heart mitochondria in two biochemically defined states

Cited by 214 publications

References 72 publications

Specific features and assembly of the plant mitochondrial complex I revealed by cryo-EM

Specific features and assembly of the plant mitochondrial complex I revealed by cryo-EM

Balanced internal hydration discriminates substrate binding to respiratory complex I

Exploring the quinone/inhibitor-binding pocket in mitochondrial respiratory complex I by chemical biology approaches

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