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Respiratory complex I

Respiratory complex I, also known as NADH:ubiquinone oxidoreductase or NADH dehydrogenase, is the first and largest enzyme complex in the mitochondrial electron transport chain (ETC), playing a pivotal role in oxidative phosphorylation.^[1] It catalyzes the oxidation of NADH derived from the tricarboxylic acid cycle, transferring two electrons to ubiquinone (coenzyme Q) to form ubiquinol, while simultaneously pumping four protons from the mitochondrial matrix to the intermembrane space per NADH oxidized.^[2] This redox reaction is coupled to proton translocation, establishing a proton motive force across the inner mitochondrial membrane that drives ATP synthesis via ATP synthase, and it represents the primary entry point for electrons into the respiratory chain.^[3] Structurally, complex I adopts an L-shaped architecture, approximately 1 MDa in molecular mass, comprising a peripheral hydrophilic arm extending into the matrix and a membrane-embedded hydrophobic arm.^[1] The peripheral arm houses the NADH-binding site, a flavin mononucleotide (FMN) cofactor, and eight iron-sulfur (Fe-S) clusters that facilitate sequential electron transfer over a distance of about 50 Å to the quinone-binding site near the membrane interface.^[2] The membrane arm, in contrast, contains proton-pumping channels and is responsible for vectorial proton translocation, with the overall structure divided into four modules: N (NADH oxidation), Q (quinone reduction), and two proton-pumping modules (P_P and P_D).^[3] In mammals, the complex assembles from 45 distinct subunits—14 core subunits conserved across species for catalytic function and 31 accessory (supernumerary) subunits that support assembly, stability, and regulatory roles—encoded by both nuclear and mitochondrial DNA.^[1] Beyond energy production, complex I is a major source of reactive oxygen species (ROS) in mitochondria, particularly under reverse electron transfer conditions, influencing cellular signaling pathways such as apoptosis and calcium homeostasis.^[1] Dysfunctions, often arising from mutations in core subunits (e.g., ND1–ND6) or assembly factors, are implicated in a spectrum of human diseases, including mitochondrial encephalomyopathies like Leigh syndrome, Leber's hereditary optic neuropathy (LHON), and neurodegenerative disorders such as Parkinson's disease.^[3] These pathologies highlight complex I's critical role in cellular metabolism, with therapeutic strategies targeting its assembly and activity under investigation to mitigate oxidative stress and energy deficits.^[1]

Overview

Definition and nomenclature

Respiratory complex I, formally known as NADH:ubiquinone oxidoreductase (EC 7.1.1.2), is the first enzyme complex in the mitochondrial electron transport chain, where it catalyzes the transfer of electrons from NADH to ubiquinone while coupling this process to proton translocation across the inner mitochondrial membrane.^[4]^[5] The complex was first isolated in the early 1960s through fractionation of submitochondrial particles from bovine heart mitochondria, with Hatefi and colleagues purifying it as the DPNH-coenzyme Q reductase and designating it as Complex I within the respiratory chain assemblies.^[6]^[1] It is commonly synonymous with NADH dehydrogenase or Type I NADH dehydrogenase, referring to its multisubunit, proton-pumping nature, in contrast to Type II NADH dehydrogenases, which are single-subunit, non-proton-pumping alternatives found in some organisms and lacking the full complexity of the respiratory chain enzyme.^[7]^[8] In mammals, respiratory complex I exhibits a molecular weight of approximately 1 MDa and comprises 45 subunits.^[9]^[5]

Evolutionary conservation

Respiratory complex I traces its origins to the last universal common ancestor (LUCA), where it likely existed as a simpler enzyme with 11 protein subunits capable of NADH oxidation and quinone reduction, as evidenced by phylogenetic analyses of archaeal and bacterial homologs.^[10] This ancestral form evolved through modular assembly from [NiFe] hydrogenases and Na+/H+ antiporter complexes, adapting to varying geochemical environments in early Earth.^[11] Homologs of complex I are widely distributed across the three domains of life: in bacteria, it is encoded by the nuo operon and functions as the primary NADH:quinone oxidoreductase; in archaea, similar enzymes with F420-dependent variants are present in lineages like Crenarchaeota; and in eukaryotes, it resides in mitochondria, reflecting endosymbiotic inheritance from α-proteobacterial ancestors.^[12]^[10] The core functional unit of complex I consists of 14 highly conserved subunits that span bacteria to mammals, encompassing the peripheral arm for electron transfer (NADH to FMN and Fe-S clusters) and the membrane arm for proton translocation.^[13] In bacteria, these 14 subunits form the minimal, fully active enzyme, sufficient for catalysis without accessory proteins.^[14] Eukaryotic versions, particularly in mammals, expand to over 44 subunits through the addition of nuclear-encoded accessory proteins that enhance stability, assembly, and regulation, though the catalytic core remains invariant.^[15] Evolutionary adaptations have led to variations in complex I across lineages, including its complete loss or reduction in certain parasitic organisms adapted to anaerobic or low-oxygen environments, such as the kinetoplastid Trypanosoma brucei, where its presence is atypical and debated in bloodstream forms.^[16] Conversely, in photosynthetic eukaryotes, a homolog known as the NAD(P)H dehydrogenase (NDH) complex persists in chloroplasts, derived from cyanobacterial ancestors and functioning in cyclic electron transport around photosystem I to optimize photoprotection and ATP production.^[17] These adaptations highlight complex I's plasticity, balancing energy demands with environmental constraints. Recent advances in cryo-electron microscopy (cryo-EM), particularly since the 2010s, have illuminated the structural conservation of complex I, revealing a canonical L-shaped architecture preserved from bacterial minimal forms to elaborate mammalian assemblies.^[18] High-resolution structures, such as those of ovine mitochondrial complex I at 3.9 Å (2016), bacterial enzymes like Thermus thermophilus at near-atomic resolution, and more recent in situ porcine structures at ~2.5–3 Å (2024), demonstrate that the L-shape—comprising a matrix-facing peripheral arm and a membrane-embedded arm—underpins electron-proton coupling across species, with only peripheral additions in eukaryotes altering the overall scaffold minimally.^[19]^[20]^[21]

Structure and assembly

Subunit composition and cofactors

Respiratory complex I exhibits a modular subunit composition that varies between prokaryotes and eukaryotes, reflecting differences in complexity and regulatory needs. In bacterial species such as Thermus thermophilus and Escherichia coli, the enzyme assumes a minimalistic form with 14 conserved core subunits—seven in the peripheral (matrix-facing) arm and seven in the membrane arm—sufficient for NADH oxidation, electron transfer to ubiquinone, and proton pumping across the membrane. These core subunits alone enable the full catalytic activity observed in simpler organisms.^[22] In mammalian mitochondria, complex I is substantially larger and more intricate, consisting of 45 subunits with a molecular mass exceeding 1 MDa. Of these, seven are encoded by mitochondrial DNA (MT-ND1, MT-ND2, MT-ND3, MT-ND4, MT-ND4L, MT-ND5, and MT-ND6), primarily integrating into the membrane arm, while the remaining 38 are nuclear-encoded and imported into the mitochondrion. The 14 core subunits, homologous to the bacterial ones, handle the primary redox and proton translocation functions, with examples including NDUFV1 (51 kDa subunit binding FMN) and NDUFS7 (hosting the terminal Fe-S cluster). The additional 31 accessory subunits, absent in bacteria, are eukaryotic innovations that enhance structural stability, facilitate stepwise assembly, and enable regulatory control; notable examples include NDUFA1–NDUFA13, which form part of early assembly intermediates and shield the complex from proteolysis or oxidative damage. Subunits like NDUFS4 stabilize the peripheral arm, NDUFA5 reinforces the Q-module for ubiquinone binding, and NDUFA12 aids in late-stage maturation by promoting cofactor integration. Regulatory accessory subunits, such as NDUFA9 (modulating ubiquinone access) and NDUFA13 (linking complex integrity to apoptotic signaling via STAT3 interaction), allow fine-tuned responses to cellular energy demands.^[1]^[22]^[15] The catalytic prowess of complex I depends on an array of non-protein cofactors embedded within the core subunits. A single flavin mononucleotide (FMN) cofactor per complex is non-covalently bound to the NDUFV1 subunit in the peripheral arm's N-module, serving as the initial electron acceptor from NADH during oxidation. Electrons are subsequently transferred through a chain of eight iron-sulfur (Fe-S) clusters—comprising two binuclear [2Fe-2S] and six tetranuclear [4Fe-4S] centers—spanning approximately 50 Å from FMN to the ubiquinone reduction site; the terminal N2 cluster, a [4Fe-4S] center in NDUFS7, is particularly critical for efficient electron donation to ubiquinone at the membrane interface. Unlike other respiratory complexes, complex I contains no heme groups, relying solely on FMN and Fe-S clusters for redox chemistry.^[19]^[22] Post-translational modifications (PTMs) on complex I subunits provide additional layers of regulation, influencing activity, assembly, and integration into supercomplexes. Phosphorylation, mediated by mitochondrial kinases such as protein kinase A or Src family members, has been documented on several core and accessory subunits; for example, the core subunit NDUFS2 (also termed 49 kDa or B14.5a) undergoes phosphorylation in bovine heart mitochondria, potentially at sites near the ubiquinone-binding interface to modulate electron transfer efficiency or conformational changes. Such modifications, often responsive to cellular signals like cAMP levels or calcium flux, underscore the dynamic regulation of complex I beyond its static composition.^[23]^[24]

Three-dimensional architecture

Respiratory complex I exhibits a characteristic L-shaped architecture, consisting of a hydrophilic peripheral arm projecting into the mitochondrial matrix and a hydrophobic membrane arm embedded in the inner mitochondrial membrane. The peripheral arm, approximately 20 nm in length, houses the flavin mononucleotide (FMN) and iron-sulfur (Fe-S) clusters essential for electron transfer, while the membrane arm, spanning about 15 nm, accommodates the proton translocation channels.^[25]^[26] The enzyme is organized into four main functional modules: the N-module, located at the tip of the peripheral arm, which binds NADH and contains FMN; the Q-module, positioned at the junction of the two arms, responsible for ubiquinone binding and reduction; and the PP (proximal) and PD (distal) modules, comprising the membrane arm, which facilitate proton pumping across the membrane.^[27] These modules are formed by the conserved core subunits, with the N- and Q-modules in the peripheral arm and the PP and PD modules in the membrane arm. Recent cryo-electron microscopy (cryo-EM) studies have resolved the structure at near-atomic resolution below 3 Å, enabling detailed visualization of subunit arrangements and cofactor positions, such as the 2.1 Å structure from Yarrowia lipolytica.^[27]^[19] Intersubunit interfaces stabilize the modular assembly, with key contacts between the peripheral and membrane arms mediated by amphipathic helices and conserved loops, such as those from ND1 interacting with NDUFS2 in the Q-module. Flexible linkers, including loops in subunits like ND1 (between transmembrane helices 5 and 6), connect the modules and permit relative movements while maintaining overall integrity. These interfaces are lined by lipids that further reinforce the structure.^[27]^[18] While the core architecture is highly conserved across species, mammalian complex I incorporates additional accessory subunits—totaling 45 compared to 14 in minimal bacterial versions—forming a stabilizing shell around the core and extending the peripheral arm slightly. Bacterial structures, such as from Escherichia coli or Thermus thermophilus, lack these supernumerary subunits, resulting in a more compact assembly with fewer lipid-binding sites and subtle variations in hydration at proton channels, like in the ND5 subunit.^[28]^[19]

Biosynthesis and assembly

The biosynthesis of respiratory complex I, also known as NADH:ubiquinone oxidoreductase, involves the coordinated integration of subunits encoded by both mitochondrial and nuclear genomes. In humans, the enzyme consists of 45 subunits, with seven hydrophobic core subunits (MT-ND1 through MT-ND6 and MT-ND4L) translated from mitochondrial DNA within the organelle, while the remaining 38 hydrophilic and accessory subunits are synthesized in the cytosol from nuclear DNA and subsequently imported into the mitochondria via translocase complexes in the outer and inner membranes.^[29]^[1] This dual-genetic origin necessitates precise temporal and spatial regulation to ensure stoichiometric balance and prevent aggregation of unassembled components. Assembly proceeds through a modular, stepwise process in the mitochondrial matrix and inner membrane, facilitated by at least 18 dedicated chaperone-like assembly factors, including the NDUFAF1 through NDUFAF12 proteins and others such as TIMMDC1 and TMEM70. These factors promote the formation of transient subcomplexes, beginning with the Q-module (comprising nuclear-encoded subunits like NDUFS2 and NDUFS3, responsible for quinone reduction), followed by integration of the N-module (nuclear-encoded subunits like NDUFV1 and NDUFS1 for NADH oxidation) and the PP and PD modules (incorporating mitochondrial-encoded subunits like MT-ND5 in the PD module for proton pumping).^[27] Recent studies have identified additional late-stage factors, such as RTN4IP1, which stabilize the ND5-module and enable docking of the ND4-module to complete the membrane arm.^[30]^[1]^[31] Key rate-limiting steps include the biogenesis and insertion of iron-sulfur (Fe-S) clusters, which are essential for electron transfer across the eight clusters in the enzyme; this process relies on the mitochondrial iron-sulfur cluster (ISC) assembly machinery, with factors like NUBPL ensuring proper maturation of the N- and Q-modules. Mitochondrial translation of the core membrane subunits represents another bottleneck, as delays can stall subcomplex progression. The entire holoenzyme matures within the inner mitochondrial membrane, where assembly factors dissociate upon completion, allowing incorporation into respiratory supercomplexes with complexes III and IV.^[30]^[31] Deficiencies in this assembly pathway, often arising from mutations in nuclear-encoded assembly factors such as NDUFAF5 or NDUFAF7, lead to isolated complex I defects characterized by reduced enzyme activity and accumulation of immature subcomplexes, contributing to mitochondrial oxidative phosphorylation disorders without affecting other respiratory complexes.^[30]^[1]

Function

Role in electron transport chain

Respiratory complex I serves as the primary entry point for electrons derived from NADH, which is produced during the tricarboxylic acid (TCA) cycle in the mitochondrial matrix. It catalyzes the oxidation of NADH to NAD⁺, transferring the two electrons along an internal chain of iron-sulfur clusters to the lipid-soluble electron acceptor ubiquinone (coenzyme Q), reducing it to ubiquinol (QH₂).^[1]^[2] This process initiates the electron transport chain (ETC) in mitochondria, linking catabolic metabolism to oxidative phosphorylation.^[27] The electron transfer in complex I is tightly coupled to the translocation of four protons (H⁺) from the mitochondrial matrix to the intermembrane space per two electrons transferred, generating a significant portion of the proton motive force (PMF) across the inner mitochondrial membrane.^[32] This contributes approximately 40% of the total proton flux in the ETC, which drives ATP synthesis by ATP synthase (complex V).^[26] Complex I integrates with downstream ETC components—complexes II, III, and IV—where electrons from QH₂ continue through the chain to reduce oxygen to water, while the PMF powers ATP production.^[27] In certain physiological contexts, such as high-intensity exercise in mammalian skeletal muscle, alternative metabolic bypasses can enable NADH oxidation independently of complex I, shunting electrons directly to complex II or other pathways to maintain energy production under stress.^[33] Overall, the complete oxidation of NADH via complex I yields approximately 2.5 molecules of ATP per NADH, compared to about 1.5 ATP from FADH₂ oxidation entering at complex II, highlighting complex I's central role in maximizing energy efficiency from TCA cycle substrates.^[34]

Overall catalytic reaction

The overall catalytic reaction of respiratory complex I (NADH:ubiquinone oxidoreductase) oxidizes NADH to NAD⁺ while reducing ubiquinone (Q) to ubiquinol (QH₂), coupling this redox process to the vectorial translocation of protons from the mitochondrial matrix to the intermembrane space. The net balanced equation is:

\text{NADH} + 5\text{H}^+_{\text{matrix}} + \text{Q} \to \text{NAD}^+ + \text{QH}_2 + 4\text{H}^+_{\text{intermembrane}}

This reflects the consumption of five protons from the matrix— one from NADH deprotonation, two for QH₂ formation, and two additional equivalents effectively pumped outward—resulting in a net translocation of four protons per reaction cycle.^[35] The stoichiometry involves the transfer of two electrons from NADH to Q, fully reducing the quinone to QH₂ without involvement of oxygen as an alternative acceptor under physiological conditions. Respiratory complex I employs a linear chain of eight iron-sulfur (Fe-S) clusters, all of the low-potential, non-Rieske type (with midpoint potentials ranging from approximately -400 mV to -150 mV), which facilitates efficient forward electron flow and minimizes superoxide formation during normoxia by limiting electron dwell times on reactive sites.^[36] In contrast to high-potential Fe-S centers like the Rieske protein in complex III, these clusters' negative potentials reduce the thermodynamic favorability of partial reduction of O₂ to superoxide in the forward direction.^[37] The reaction is thermodynamically favorable, driven by a substantial redox potential span: the NADH/NAD⁺ couple has a midpoint potential (E_m) of -320 mV, while the Q/QH₂ couple operates at approximately +90 mV, yielding a ΔE of about 410 mV and releasing sufficient free energy (ΔG°' ≈ -79 kJ/mol) to support proton pumping.^[38] The flavin mononucleotide (FMN) cofactor, with an E_m near -340 mV, serves as the initial electron acceptor from NADH, bridging to the Fe-S chain.^[39] Experimental verification of the reaction kinetics and stoichiometry has relied on stopped-flow spectroscopy, which captures millisecond-scale electron transfer events by monitoring absorbance changes during NADH oxidation and Q reduction in real time. These studies confirm the rapid, sequential reduction of FMN and Fe-S clusters, with overall turnover rates supporting the 4 H⁺/2 e⁻ stoichiometry in both bacterial and mammalian enzymes under physiological proton motive force.^[40]^[32]

Mechanism

Electron transfer pathway

The electron transfer pathway in respiratory complex I initiates with the binding of NADH at the peripheral arm, where it undergoes oxidation by transferring a hydride ion to the non-covalently bound flavin mononucleotide (FMN) cofactor, forming NAD⁺ and FMNH₂. This step occurs rapidly through a proton-coupled hydride transfer mechanism, enabling efficient entry of electrons into the complex.^[41] The FMN is embedded within the NDUFV1 subunit and positioned to accept electrons without direct involvement of protein residues in the initial redox event.^[42] From FMNH₂, the two electrons are relayed sequentially in one-electron steps through a chain of eight iron-sulfur (Fe-S) clusters spanning over 95 Å from the FMN to the ubiquinone (Q)-binding site at the interface of the peripheral and membrane arms. The primary linear pathway proceeds as FMN → N3 → N1b → N4 → N5 → N6a → N6b → N2 → Q, with electron tunneling facilitating transfers across cluster-to-cluster distances of 10–14 Å.^[43] These transfers are enhanced by intervening water molecules and protein residues, such as cysteines, which bridge gaps and increase rates by up to three orders of magnitude in key steps like N5 to N6a.^[43] An off-pathway [2Fe-2S] cluster (N1a) near FMN can occasionally participate in reverse transfers under certain conditions but does not contribute to forward flux.^[41] The Fe-S clusters comprise two binuclear [2Fe-2S] types (N1a and N1b) and six tetranuclear [4Fe-4S] types (N2, N3, N4, N5, N6a, N6b), coordinated primarily by cysteine residues within subunits like NDUFS1, NDUFS7, and NDUFS8.^[42] The terminal [4Fe-4S] cluster N2, located in the NDUFS2 subunit approximately 12 Å from the Q site, acts as a gatekeeper by delivering electrons directly to the ubiquinone headgroup, which binds in a narrow tunnel involving conserved residues like Tyr87 and His38.^[42] This positioning ensures controlled access and reduction of Q to semiquinone and then quinol.^[44] Redox potentials are tuned progressively higher along the chain to drive forward transfer and minimize reversal, starting at approximately -380 mV for FMN and N3, rising through intermediate clusters (N1b ≈ -250 mV, N4/N5 ≈ -250 to -300 mV), to -150 mV for N2, and culminating at +90 mV for the Q/QH₂ couple.^[42] Recent molecular dynamics simulations from the 2020s indicate that the final transfer from N2 to Q involves a coordinated, potentially bifurcated delivery of the two electrons required for full Q reduction, possibly via a stabilized semiquinone intermediate within the Q tunnel to optimize energetics and coupling.^[14]

Proton translocation process

Respiratory complex I translocates four protons across the inner mitochondrial membrane for every two electrons transferred from NADH to ubiquinone, establishing a proton motive force essential for ATP synthesis.^[32] This process operates independently of a Q-cycle, unlike complexes III and IV, relying instead on direct coupling between quinone reduction and vectorial proton pumping through dedicated membrane channels.^[13] The mechanism involves conformational waves initiated at the quinone-binding site (Q-site), which propagate through the peripheral arm to the membrane domain, coordinating proton uptake from the matrix and release to the intermembrane space.^[13] Proton translocation occurs via four distinct pumping channels, three of which are formed by the antiporter-like subunits ND2, ND4, and ND5, while the fourth is the E-channel primarily involving ND1.^[45] Each channel features half-channel architectures: N-side half-channels for proton uptake from the matrix, connected by central binding sites, and P-side half-channels for release to the intermembrane space.^[45] In ND2, ND4, and ND5, key residues such as conserved glutamates and lysines (e.g., Glu in ND4) facilitate proton conduction along hydrated pathways, with water wires enabling Grotthuss-like transfer; the Q-entry path links to the quinone tunnel near ND1 and ND3, while H+ exit paths converge toward ND5 for coordinated ejection.^[45] These channels ensure unidirectionality, preventing backflow under the electrochemical gradient.^[13] The energy for uphill proton pumping derives from electrostatic repulsion generated during ubiquinol (QH₂) formation and deprotonation at the Q-site, which triggers a cascade of charged-group rearrangements propagating as an electrostatic wave through the complex.^[46] This repulsion displaces ionizable residues and helix bundles in the membrane arm, driving protonation changes in the channels against the proton motive force.^[46] Experimental support for this mechanism comes from site-directed mutagenesis studies, where introducing disulfide bonds between the ND3 TMH1-2 loop and NDUFS7 subunit decouples conformational changes from proton pumping, abolishing translocation without affecting electron transfer.^[13] Additionally, pH-jump assays combined with molecular dynamics simulations demonstrate rapid water chain reorganization in the channels upon redox-state changes, confirming dynamic proton pathways responsive to Q-site events.^[13] High-resolution cryo-EM structures further validate the half-channel geometries and key residues, showing hydrated networks essential for conduction.^[45]

Conformational dynamics

Respiratory complex I exhibits dynamic conformational changes throughout its catalytic cycle, enabling the coupling of electron transfer from NADH to ubiquinone reduction with proton translocation across the membrane. These dynamics primarily involve transitions between distinct structural states at the quinone (Q)-binding site, captured through high-resolution cryo-electron microscopy (cryo-EM) structures. The enzyme alternates between an open state, which facilitates ubiquinone access and semiquinone release, and a closed state that seals the Q-site to support stable reduction and prevent uncoupled activity.^[45] These state transitions occur at the interface between the peripheral (NADH-oxidizing) arm and the membrane arm, involving coordinated rearrangements of loops and helices in core subunits such as NDUFS2, ND1, and ND3.^[47] Ubiquinone binding to the Q-site triggers a key conformational rearrangement, including a rotation of the peripheral arm by approximately 10° relative to the membrane arm, which constricts the Q-cavity and positions the substrate for reduction.^[14] This motion, observed in bacterial and mammalian structures, tightens interactions around the quinone headgroup, stabilizing the semiquinone intermediate and initiating downstream signaling. Cryo-EM intermediates from 2018 onward, including turnover states in ovine and human complex I, reveal open Q-sites with disordered access loops and closed configurations with ordered, solvent-excluded channels, highlighting the precision of these changes during catalysis.^[48]^[47] The conformational dynamics propagate as waves of structural rearrangement from the Q-module (encompassing the FMN and iron-sulfur clusters) to the distal P-module (proton-pumping channels), synchronizing electron transfer with proton release into four half-channels.^[13] These waves, inferred from comparative structural analyses and molecular dynamics simulations, ensure mechanical coupling without energy dissipation, with local twisting and tilting motions amplifying the signal across the ~200 Å span of the membrane arm. Under conditions of low proton motive force (ΔpH), the closed state predominates to inhibit reverse electron flow, blocking futile cycling and superoxide generation at the Q-site.^[49]

Regulation and modulation

Active/inactive transitions

Respiratory complex I in mammalian mitochondria exists in two interconvertible states: an active (A) form that catalyzes rapid NADH oxidation and a deactive (D) form that is catalytically inactive and predominant in isolated mitochondria lacking substrates. The D-form arises spontaneously during hypoxia or ischemia when electron turnover ceases, serving as a protective mechanism to halt electron flow and minimize reactive oxygen species (ROS) production upon reoxygenation. Activation from the D- to A-form requires energization by the proton motive force (ΔpH) across the inner mitochondrial membrane or exposure to chaotropic agents such as urea, which disrupt the stable D-state conformation. This transition involves exposure of Cys39 in the ND3 subunit, which becomes solvent-accessible in the D-form, allowing modification by thiol-reactive agents.^[50] The physiological role of the A/D transition is to safeguard mitochondria during metabolic stress, such as ischemia, by suppressing ROS generation at complex I, the primary site of superoxide production under reverse electron transfer conditions. During ischemia, the D-form predominates, preventing harmful ROS bursts that could damage cellular components upon reperfusion; reactivation occurs slowly over minutes as oxygen and substrates become available, contrasting sharply with the millisecond timescale of normal catalytic cycles. This temporal mismatch ensures controlled resumption of respiration, reducing oxidative injury in tissues like the heart and brain. Studies in isolated mitochondria and cellular models confirm that enforced maintenance of the A-form exacerbates ischemia-reperfusion damage, underscoring the protective utility of the D-state. Structurally, the D-form features localized disorder in the ubiquinone-binding channel, including a small rotation (approximately 3.4°) of the hydrophilic domain relative to the membrane arm in the NDUFS1 subunit (the 75-kDa iron-sulfur protein), which displaces key elements like the β1-β2 loop of the 49-kDa subunit and prevents substrate access.^[51] Cryo-electron microscopy structures reveal that this rotation and associated unfolding block the Q-site, rendering the enzyme inactive until corrective conformational changes restore order. Recent 2022 investigations have identified thiol oxidation of conserved cysteines in NDUFS2 and adjacent subunits as a key trigger for the D-state, linking redox signaling to the transition and highlighting its role in hypoxic adaptation across species like C. elegans.^[52] These oxidation events propagate structural shifts that stabilize the D-form, reversible by reducing agents or ΔpH-driven protonation. In contrast to mammalian complex I, bacterial NADH:ubiquinone oxidoreductases lack this A/D regulatory switch, remaining constitutively active without the slow deactivation kinetics or protective D-state, a distinction attributed to the absence of certain accessory subunits and the simpler membrane environment in prokaryotes. This eukaryotic-specific mechanism likely evolved to fine-tune respiration in response to fluctuating oxygen levels and metabolic demands in multicellular organisms.

Pharmacological inhibitors

Pharmacological inhibitors of respiratory complex I target various sites within the enzyme, primarily the ubiquinone (Q)-binding channel, to block electron transfer from NADH to ubiquinone and disrupt proton pumping. These compounds have been instrumental in elucidating the enzyme's mechanism and hold potential for therapeutic applications in cancer and antiparasitic treatments.^[53]^[54] Rotenone, a natural isoflavonoid derived from plant roots, is a prototypical high-affinity inhibitor that binds within the Q-reduction site near the N2 iron-sulfur cluster, involving interactions with conserved residues such as Tyr108 and His59 in the NDUFS2 subunit. This binding sterically hinders ubiquinone access and electron transfer, with an IC50 in the low nanomolar range in bovine mitochondrial assays.^[53] Piericidin A, an α-pyridone antibiotic produced by actinomycetes, similarly occupies the Q-binding site at the tunnel's apex, blocking N2 cluster access and ubiquinone reduction, also achieving low nanomolar IC50 values; structural studies reveal cooperative binding that may involve a secondary site deeper in the Q-tunnel.^[53]^[3] ADP-ribose acts as a physiological, reversible competitive inhibitor of NADH oxidation at the FMN-binding site in the enzyme's peripheral arm, with inhibition constants in the micromolar range, potentially modulating complex I activity under oxidative stress conditions.^[55] Metformin, an antidiabetic biguanide, exerts partial inhibition through binding in the amphipathic Q-channel near the membrane interface, displacing structural elements like the ND5 helix and preferring the deactive enzyme state, though its potency is weak with IC50 values exceeding 10 mM in isolated membranes.^[56] Stigmatellin, primarily an inhibitor of complex III, shows off-target effects on complex I at higher concentrations as a class B inhibitor, noncompetitively blocking activity without direct competition at the Q-site.^[57] Cryo-EM structures of bacterial and mammalian complex I have mapped inhibitor binding to the Q-cleft, a narrow tunnel extending from the membrane domain, informing design of selective agents.^[3] In recent developments, IACS-010759 represents a potent, selective small-molecule inhibitor that binds tightly within the Q-cavity via a "cork-in-bottle" mechanism, achieving subnanomolar potency; it underwent phase I clinical trials for oncology starting in 2016 but development was halted due to toxicity concerns as of 2025.^[58]^[59] These inhibitors, particularly rotenone and piericidin analogs, also show promise in antiparasitic applications by targeting parasite complex I variants.^[54]

Reactive oxygen species

Superoxide generation mechanisms

Respiratory complex I generates superoxide primarily through the reduction of molecular oxygen by the flavin mononucleotide (FMN) cofactor at site I_F, particularly via its reduced forms, including the FMN semiquinone intermediate, which exhibits a high redox potential conducive to electron leakage. A secondary site, I_Q at the quinone-binding channel, also contributes to superoxide production, especially during forward electron transfer when the ubiquinone pool is reduced.^[60] This process occurs at the FMN site in the enzyme's hydrophilic arm, where electrons from NADH can divert off the main transfer pathway to react with O₂, forming superoxide (O₂⁻•) as the initial product, with subsequent dismutation potentially yielding hydrogen peroxide.^[36] Studies on isolated bovine complex I confirm that the terminal iron-sulfur cluster N₂ does not significantly contribute to superoxide production, reinforcing the FMN and Q sites as the dominant locations.^[60] Superoxide production is modulated by the direction of electron flow within complex I. In the forward mode, driven by NADH oxidation under normoxic conditions, superoxide generation remains low, typically accounting for approximately 1-2% of electrons diverted from the catalytic cycle to O₂.^[61] Conversely, during reverse electron transfer (RET)—facilitated by a highly reduced ubiquinone pool and elevated proton-motive force, as seen in hypoxic or ischemic states—superoxide output increases dramatically, up to several-fold higher than in forward mode, due to hyper-reduction of the FMN site. RET-driven superoxide primarily emanates from the fully reduced FMN-hydroquinone (FMNH₂), serving as a key intermediate that reacts with O₂.^[62]^[60] Kinetic analyses using pulse radiolysis have elucidated the reaction rates at the FMN site, revealing second-order rate constants for the interaction between reduced FMN species and O₂ on the order of 10³ M⁻¹ s⁻¹, which limits superoxide formation to a minor fraction of overall electron flux.^[63] These measurements highlight the FMN-hydroquinone as the reactive species, with superoxide production rates in isolated complex I reaching about 0.15% of maximal catalytic turnover under saturating NADH conditions (based on studies in Yarrowia lipolytica).^[64] Studies have shown that the mitochondrial matrix NADH/NAD⁺ ratio plays a key role in partitioning superoxide production between forward and reverse modes. Elevated NADH/NAD⁺ ratios, indicative of reductive stress, enhance FMN reduction and thereby amplify superoxide leakage during forward electron transfer, while favoring RET under high membrane potential; this dynamic positions complex I as a redox sensor responsive to metabolic state.^[65]

Physiological and pathological roles

Complex I-derived reactive oxygen species (ROS) play essential physiological roles at low levels, acting as signaling molecules that facilitate cellular adaptation and maintain redox homeostasis. In hypoxic conditions, modest ROS production from Complex I stabilizes hypoxia-inducible factor-1α (HIF-1α), promoting adaptive responses such as angiogenesis and metabolic reprogramming to enhance oxygen utilization.^[66] This signaling mechanism helps cells sense and respond to environmental stressors, ensuring survival under low-oxygen states. Additionally, low-level Complex I ROS contribute to redox homeostasis by modulating antioxidant defenses and thiol-based signaling pathways, preventing excessive oxidative damage while supporting normal mitochondrial function. Recent studies (as of 2025) have further highlighted Complex I ROS as metabolic sensors in immune cells, where they fine-tune activation and effector functions, such as in macrophages, by integrating nutrient availability with inflammatory signaling during chronic stimulation.^[67]^[68] In pathological contexts, elevated ROS from Complex I drive cellular damage and dysfunction. High ROS levels oxidize mitochondrial DNA (mtDNA), leading to mutations and impaired respiratory chain assembly, which exacerbates energy deficits in affected cells.^[69] This oxidative stress also triggers apoptosis through activation of the intrinsic mitochondrial pathway, involving cytochrome c release and caspase cascades, particularly in response to prolonged metabolic insults.^[70] Furthermore, during ischemia-reperfusion injury, burst-like Complex I ROS production via reverse electron transport contributes to tissue damage by promoting inflammation and necrosis in organs like the heart and brain.^[71] Modulation of Complex I ROS represents a therapeutic strategy to balance its dual roles, with mitochondria-targeted antioxidants like MitoQ selectively scavenging superoxide at this site to mitigate oxidative harm without fully disrupting electron transport.^[72] Evolutionarily, the propensity for Complex I to generate ROS reflects a trade-off between high respiratory efficiency and the risk of oxidative leakage, allowing rapid energy production but constraining longevity in high-metabolism organisms.^[73]

Pathological implications

Associated mitochondrial diseases

Respiratory complex I deficiency is the most common cause of mitochondrial disease in children, accounting for approximately 30% of cases, and manifests primarily as primary genetic disorders such as Leigh syndrome.^[74] Leigh syndrome, the most prevalent presentation, affects about 1 in 40,000 individuals and is linked to complex I defects in roughly 30% of cases, often due to mutations in mitochondrial DNA (mtDNA) genes encoding complex I subunits, particularly the MT-ND genes (e.g., MT-ND1, MT-ND2, MT-ND3, MT-ND4, MT-ND5, MT-ND6).^[75]^[76] Complex I deficiency accounts for approximately 30% of pediatric mitochondrial diseases, which have an overall prevalence of about 1 in 5,000 births.^[75] Clinical features typically include neonatal-onset lactic acidosis, psychomotor regression, hypotonia, seizures, and characteristic bilateral symmetric lesions in the basal ganglia and brainstem visible on neuroimaging.^[77] Other associated disorders include Leber's hereditary optic neuropathy (LHON), which primarily causes acute or subacute bilateral vision loss due to optic nerve atrophy from mutations in MT-ND1, MT-ND4, or MT-ND6 (e.g., m.11778G>A in MT-ND4), and may occasionally overlap with Leigh-like features.^[77] Some MT-ND mutations, such as those in MT-ND5, can present with MELAS (mitochondrial encephalomyopathy, lactic acidosis, and stroke-like episodes) overlap syndromes, featuring recurrent stroke-like episodes alongside lactic acidosis and myopathy.^[77] Isolated complex I-deficient myopathies are also reported, characterized by progressive muscle weakness, exercise intolerance, and elevated serum lactate, without predominant neurological involvement.^[77] Diagnosis of these complex I-related diseases relies on a combination of clinical evaluation, biochemical testing, and genetic analysis. Enzyme assays measuring NADH:ubiquinone oxidoreductase activity in skin fibroblasts, muscle, or blood leukocytes confirm reduced complex I function, often below 30% of control values.^[77] Muscle biopsy may reveal ragged-red fibers, subsarcolemmal accumulation of mitochondria, and cytochrome c oxidase (COX) deficiency on histochemistry.^[77] Definitive diagnosis involves next-generation sequencing (NGS) of mtDNA and nuclear DNA to identify pathogenic variants, such as heteroplasmic MT-ND mutations with heteroplasmy levels typically exceeding 70-80% in affected tissues.^[75]^[77] Recent advances in 2024 include preclinical gene therapy approaches using adeno-associated virus (AAV) vectors for allotopic expression of MT-ND5 in Leigh syndrome models caused by mtDNA mutations, demonstrating restored complex I assembly, improved ATP production, and extended survival in mice without significant off-target effects. As of 2025, emerging enzyme-based technologies for precise mtDNA editing in patient-derived cells show promise for correcting complex I mutations, complementing AAV gene therapy approaches.^[78]^[77]^[79] These strategies relocate the mutant mtDNA gene to the nucleus for cytosolic transcription and mitochondrial import of the protein, offering promise for monogenic mtDNA disorders, though clinical translation remains challenged by heteroplasmy and tissue-specific delivery.^[79]

Links to neurodegeneration and aging

Respiratory complex I dysfunction plays a central role in Parkinson's disease (PD) pathogenesis, particularly through its contribution to the selective degeneration of dopaminergic neurons in the substantia nigra pars compacta. Postmortem analyses of PD brains reveal a 30-40% reduction in complex I activity specifically in this region, which correlates with approximately 50-60% neuronal loss at the symptomatic onset of motor impairments.^[80]^[81] This deficiency is not uniform across the brain but is most pronounced in vulnerable dopaminergic populations, exacerbating energy deficits and promoting cell death. The toxin MPTP, which induces a parkinsonian syndrome, is converted to MPP+ that potently inhibits complex I in a manner akin to rotenone, thereby replicating the mitochondrial impairment observed in idiopathic PD and underscoring complex I as a key therapeutic target.^[82] In Alzheimer's disease (AD), amyloid-β (Aβ) oligomers inhibit the enzymatic activity of complex I, disrupting electron transport in neuronal mitochondria. This inhibition precedes plaque formation in AD mouse models and contributes to synaptic dysfunction and cognitive decline by reducing ATP production and amplifying oxidative stress. Therapeutic interventions targeting complex I, including coenzyme Q10 supplementation, have demonstrated neuroprotective effects in preclinical AD models by restoring mitochondrial function, reducing Aβ toxicity, and improving memory performance, although large-scale human trials are ongoing to validate these benefits.^[83]^[84]^[85] Aging is associated with a progressive decline in complex I activity, with reported reductions of 20-50% in various tissues, including brain and muscle, by advanced age, driven by somatic mitochondrial DNA mutations that accumulate over time and lead to impaired assembly and function. This age-related deterioration fosters a cycle of mitochondrial inefficiency, distinct from inherited deficiencies, and heightens susceptibility to neurodegenerative conditions. A 2024 multicenter study stratifying idiopathic PD patients by complex I deficiency levels identified it as a robust biomarker for disease subtypes and progression, with severe deficiencies predicting non-tremor-dominant phenotypes and faster decline, informing potential monitoring strategies in aging populations.^[86]^[87]

Occurrence in other organisms

Bacterial NADH dehydrogenases

Bacterial NADH dehydrogenases, also known as NDH-1 or type I NADH:quinone oxidoreductases, serve as prokaryotic homologs of mitochondrial respiratory complex I, embedded in the plasma membrane to facilitate energy conservation.^[26] These enzymes are encoded by the conserved nuo operon, which typically comprises 14 core subunits designated NuoA through NuoN, forming a ~500 kDa L-shaped complex with a peripheral arm for electron transfer and a membrane arm for proton translocation.^[88] Prominent examples include the complexes from Escherichia coli and Thermus thermophilus, which have been extensively studied as minimal models due to their structural simplicity and genetic tractability. Functionally, bacterial NADH dehydrogenases catalyze the transfer of electrons from NADH to quinone (ubiquinone or menaquinone, depending on the species), coupled to proton pumping across the membrane, generating a proton motive force for ATP synthesis.^[35] The proton-to-electron stoichiometry varies but is commonly 4 H⁺ per 2 e⁻, enabling efficient energy coupling in aerobic respiration; however, some variants exhibit 2–3 H⁺/2 e⁻, reflecting adaptations to environmental conditions.^[35] In facultative anaerobes like Rhodobacter sphaeroides, these enzymes also support anaerobic respiration by linking NADH oxidation to alternative electron acceptors such as fumarate, highlighting their versatility in diverse metabolic contexts.^[89] Structural studies have provided key mechanistic insights, with the first atomic model of the complete bacterial complex from T. thermophilus determined in 2010 at 4.5 Å resolution using X-ray crystallography, revealing 63 transmembrane helices in the membrane arm and conserved iron-sulfur clusters in the peripheral arm.^[90] Unlike eukaryotic complex I, bacterial versions lack accessory (supernumerary) subunits, resulting in a more streamlined architecture and simpler regulation without the extensive post-translational modifications or assembly factors seen in mitochondria.^[26] This minimal design has made bacterial complexes ideal for biophysical and mechanistic investigations, elucidating how long-range conformational changes propagate from the quinone-binding site to drive proton pumping through antiporter-like subunits (NuoL, NuoM, NuoN).^[90] Beyond fundamental biology, bacterial NADH dehydrogenases represent promising therapeutic targets, particularly in pathogenic species like Mycobacterium tuberculosis, where complex I is essential for survival under hypoxia and oxidative stress, and inhibitors disrupting its activity show potential as novel antibiotics against drug-resistant strains.^[91]

Chloroplast NDH complexes

The chloroplast NAD(P)H dehydrogenase-like (NDH) complex is a large multi-subunit enzyme embedded in the thylakoid membranes of higher plant chloroplasts, serving as a ferredoxin:plastoquinone reductase that facilitates cyclic electron flow around photosystem I (PSI).^[92] This process recycles electrons from ferredoxin back to plastoquinone, bypassing photosystem II and thereby enhancing ATP production without net NADPH synthesis, which supports photosynthetic efficiency under varying light conditions.^[93] The complex's activity is crucial for maintaining redox balance in the photosynthetic electron transport chain, particularly during high-light stress when linear electron flow may generate excess reducing power.^[94] Composed of approximately 29 subunits organized into five subcomplexes, the NDH complex includes 11 plastid-encoded subunits (NdhA through NdhK, homologous to bacterial NDH-1 components) and at least 18 nuclear-encoded subunits unique to plants, such as NDH-M, -N, and -O, which are essential for assembly and function.^[95] These nuclear-encoded subunits enable specific adaptations, including interactions with PSI light-harvesting complexes.^[96] In contrast to mitochondrial complex I, which pumps four protons per two electrons, the chloroplast NDH complex exhibits a more limited proton-pumping capacity of about two protons per two electrons, prioritizing photoprotection through ΔpH generation for non-photochemical quenching over maximal energy conservation.^[97] This reduced pumping aligns with its role in stromal acidification to regulate electron flow and prevent over-reduction of the plastoquinone pool.^[98] The NDH complex performs several key functions beyond cyclic electron transport, including support for CO₂ concentration mechanisms in C₄ plants, where it sustains a high ATP/ADP ratio in bundle sheath cells to enhance carbon fixation efficiency. It also contributes to reactive oxygen species (ROS) scavenging by oxidizing excess reductants and mitigating photooxidative damage, as evidenced by increased ROS accumulation and photosynthetic impairment in NDH-deficient tobacco mutants under high light.^[99] Notably, the NDH complex is absent in the plastids of most eukaryotic algae, with ndh genes lost from all sequenced algal plastid genomes except those of Charophyceae and certain Prasinophyceae, reflecting evolutionary dispensability in non-vascular photosynthetic organisms.^[100] Recent cryo-electron microscopy (cryo-EM) structures from 2022 have elucidated the architecture of the NDH complex as part of a hybrid PSI-NDH supercomplex in barley (Hordeum vulgare), revealing how up to two PSI cores associate with one NDH unit via stromal-facing protrusions, stabilized by shared lipids and chlorophylls for efficient electron transfer.^[101] These structures highlight plant-specific subunits that facilitate supercomplex formation, underscoring the NDH's integration into the photosynthetic apparatus for coordinated cyclic flow. A more recent 2025 cryo-EM structure of the NDH–PSI–LHCI supercomplex from spinach (Spinacia oleracea) at 3.0–3.3 Å resolution further details the assembly, consisting of 41 protein subunits reinforced by 46 lipids, providing additional insights into subunit interactions and lipid mediation.^[102]

Genetics

Encoding genes in humans

In humans, respiratory complex I (NADH:ubiquinone oxidoreductase) is encoded by a combination of mitochondrial and nuclear genes, reflecting its dual-genome origin. The seven core subunits derived from mitochondrial DNA (mtDNA) are essential components of the membrane arm, while the 38 nuclear-encoded subunits include both core and accessory proteins that form the peripheral and membrane arms, as well as structural and regulatory elements. These 38 subunits are produced from 37 autosomal genes, as the NDUFAB1 gene encodes two copies of its acyl carrier protein-like subunit. The mtDNA-encoded genes are MT-ND1, MT-ND2, MT-ND3, MT-ND4, MT-ND4L, MT-ND5, and MT-ND6. These genes reside on the circular, maternally inherited mtDNA molecule and produce proteins synthesized directly within the mitochondrial matrix. Transcription of these genes occurs as part of long polycistronic transcripts from the heavy and light strands of mtDNA, which are processed into mature mRNAs for the 13 mtDNA-encoded respiratory chain proteins, including the seven for complex I.^[1] The nuclear genome encodes the majority of complex I subunits via 37 autosomal genes, whose products are translated in the cytosol and subsequently imported into mitochondria via specific targeting signals. These subunits are organized into functional families based on their roles in electron transfer, proton pumping, and structural stability. For instance, the iron-sulfur (Fe-S) cluster-containing subunits are encoded by NDUFS1 through NDUFS8, which harbor the redox-active Fe-S centers critical for electron transfer from NADH. Accessory subunits, often involved in assembly and stability, include those from the NDUFA1 through NDUFA13 family, such as NDUFA1 (MWFE) and NDUFA13 (GRIM-19), which contribute to the matrix-facing arm. Other families encompass NDUFV1 and NDUFV2 (flavin mononucleotide-binding subunits), NDUFB1 through NDUFB11 (membrane arm accessories), NDUFC1 and NDUFC2 (core membrane components), and NDUFAB1 (acyl carrier protein-like subunit, with two copies). This nuclear contribution ensures the complex's ~980 kDa mass and L-shaped architecture.^[103]^[104]^[1] Expression of the nuclear-encoded complex I genes is tightly coordinated by the transcriptional coactivator peroxisome proliferator-activated receptor gamma coactivator 1-alpha (PGC-1α), which integrates environmental signals to drive mitochondrial biogenesis. PGC-1α interacts with nuclear respiratory factors (NRF1 and NRF2) and estrogen-related receptors (ERRα/γ) to upregulate these genes, ensuring balanced synthesis with mtDNA-encoded subunits.^[105] In contrast, mtDNA transcription is regulated independently by mitochondrial transcription factor A (TFAM), though PGC-1α indirectly influences it via NRF1-mediated TFAM expression. The MitoCarta database catalogs over 1,100 human mitochondrial proteins, including all complex I subunits, with annotations for their subcellular localization, functional domains, and expression patterns based on mass spectrometry and GFP-tagging data. This resource facilitates identification and validation of encoding genes in mitochondrial proteomics studies.

Mutations and genetic variants

Mutations in genes encoding subunits or assembly factors of respiratory complex I are a leading cause of isolated complex I deficiency, accounting for approximately 25% of mitochondrial respiratory chain disorders in children. These mutations can occur in either mitochondrial DNA (mtDNA) or nuclear DNA (nDNA), with mtDNA variants exhibiting heteroplasmy—the coexistence of mutant and wild-type mtDNA molecules—where disease manifestation typically requires a mutant load exceeding 70% in affected tissues due to the threshold effect. Common mutation types include missense variants that alter amino acid sequences, often disrupting iron-sulfur (Fe-S) cluster coordination essential for electron transfer, and frameshift or nonsense mutations leading to premature termination and truncated proteins that impair complex stability.^[76]^[106] A well-documented mtDNA variant is the m.13513G>A missense mutation in MT-ND5, which substitutes aspartic acid for asparagine (p.D393N) in the ND5 subunit, frequently associated with Leigh syndrome and characterized by variable heteroplasmy levels correlating with clinical severity. In nDNA, mutations in NDUFS1, encoding the 75 kDa Fe-S subunit, are prevalent; for instance, the homozygous c.1222C>T (p.R408C) missense mutation has been identified in an Asian child with Leigh-like encephalopathy, resulting in profoundly reduced complex I activity in fibroblasts and brain tissue. These examples highlight how specific variants target critical structural domains, such as the Q-module in ND5 or Fe-S binding sites in NDUFS1.^[107]^[108] The molecular consequences of these mutations often involve defective complex I assembly, leading to accumulation of subcomplexes and decreased holoenzyme formation, as seen in NDUFS1 variants where Fe-S cluster instability reduces NADH oxidation efficiency. Frameshift mutations, such as those introducing premature stop codons, exacerbate this by promoting proteasomal degradation of nascent subunits, further destabilizing the complex. Additionally, mutant complex I generates elevated reactive oxygen species (ROS) due to electron leakage from disrupted Fe-S centers, contributing to oxidative damage in high-energy tissues like the brain. Genotype-phenotype correlations reveal that mtDNA mutations like m.13513G>A tend to present with encephalomyopathy at lower heteroplasmy thresholds in neural cells (~60-80%), while severe nDNA biallelic variants, such as in NDUFS1, often cause earlier-onset, rapidly progressive disease with poor prognosis.^[76]^[109] Recent studies have advanced understanding of assembly defects, with biallelic loss-of-function mutations in NDUFAF2, an essential chaperone for the N-module attachment during complex I biogenesis, reported in 2024 as causing a severe infantile brainstem-predominant neurodegenerative disorder akin to Leigh syndrome, underscoring the role of assembly factors in pathogenesis. Experimental approaches, including CRISPR/Cas9-mediated knockout models of complex I genes like NDUFS4 in induced pluripotent stem cells, have recapitulated assembly failures and ROS overproduction, providing platforms to test therapeutic interventions for these variants. Such mutations link to mitochondrial diseases like Leigh syndrome, where complex I deficiency manifests as subacute necrotizing encephalomyelopathy.^[110]^[111]

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Different Functions of Phylogenetically Distinct Bacterial Complex I ...
Complex I activity is required for anaerobic growth of R. sphaeroides. The R. sphaeroides genome has two operons that encode complex I homologues (Fig. 1A). One ...
[80]
The architecture of respiratory complex I - Nature
The architecture of respiratory complex I. Download PDF. Article; Published: 27 May 2010. The architecture of respiratory complex I. Rouslan G. Efremov, ...
[81]
Structure of mycobacterial respiratory complex I - PNAS
The core Complex I structure consists of 14 subunits, with 31 supernumerary subunits in mammalian mitochondria (28, 34), and total molecular masses ranging from ...Sign Up For Pnas Alerts · Results · Active Mycobacterial Complex...
[82]
An Src Homology 3 Domain-Like Fold Protein Forms a Ferredoxin ...
This study indicates that CRR31, a novel NDH subunit, serves in the high-affinity binding of ferredoxin to NDH and proposes that chloroplast NDH is ferredoxin: ...
[83]
The Significance of Chloroplast NAD(P)H Dehydrogenase Complex ...
Apr 22, 2021 · The significance of the NDH-mediated cyclic electron transport is discussed from the perspective of plant evolution and physiological ...
[84]
Chloroplast NDH: A different enzyme with a structure similar to that ...
Structural evolution of photosynthetic NDH. Chloroplast NDH originated from the cyanobacterial NDH-1 complex. In Synechocystis PCC6803, NDH-1 is involved in the ...
[85]
Supramolecular assembly of chloroplast NADH dehydrogenase-like ...
Mar 7, 2022 · In angiosperms, the NDH complex is composed of at least 29 subunits, which can be divided into five subcomplexes (Ifuku et al., 2011; Shikanai, ...
[86]
Evolution of an assembly factor-based subunit contributed to a novel ...
Jun 17, 2021 · In contrast, the molecular size of the chloroplast NDH complex has gradually increased during the evolution of land plants. Our results ...
[87]
The higher plant plastid NAD(P)H dehydrogenase-like complex ...
We demonstrate that the chloroplast NADPH dehydrogenase complex, a homolog to respiratory Complex I, pumps approximately two protons from the chloroplast stroma ...
[88]
Chloroplast NADH dehydrogenase‐like complex‐mediated cyclic ...
Jul 22, 2024 · During the evolutionary transition from C3 to C4 plants, NDH levels increased concurrently with the emergence of the C4 cycle as a CO2 pump (Lyu ...
[89]
Chloroplastic NAD(P)H Dehydrogenase in Tobacco Leaves ...
However, there was no direct evidence for involvement of the NDH pathway in ROS scavenging. This work provides direct evidence for the involvement of the ...
[90]
Plastid ndh genes in plant evolution - ScienceDirect.com
The ndh genes are absent in all sequenced plastid DNAs of algae except for the Charophyceae and some Prasinophyceae.
[91]
The nuclear encoded subunits of complex I from bovine heart ...
The sequences of 45 subunits of complex I have been determined. Seven of them are encoded by mitochondrial DNA, and 38 are products of the nuclear genome.
[92]
Human CIA30 is involved in the early assembly of mitochondrial ...
Of the complex I subunits, seven are encoded by mitochondrial DNA (mtDNA) whereas the remainder are encoded by nuclear genes, translated in the cytosol and ...
[93]
Errα and Gabpa/b specify PGC-1α-dependent oxidative ... - PNAS
Fig. 2. Proposed model of mechanism of action of PGC-1α. PGC-1α is a highly regulated gene that responds to external stimuli, e.g., reduced in diabetes and ...
[94]
Mitochondrial DNA heteroplasmy in disease and targeted nuclease ...
Feb 19, 2020 · This threshold is dependent on the mutation, the cell type, and tissue type, and can vary between 60 and 90% mutant mtDNA for an detectable ...
[95]
The mitochondrial 13513G > A mutation is most frequent in Leigh ...
The mitochondrial 13513G > A mutation is most frequent in Leigh syndrome combined with reduced complex I activity, optic atrophy and/or Wolff-Parkinson-White.Missing: 12308A> | Show results with:12308A>
[96]
The p.M292T NDUFS2 mutation causes complex I-deficient Leigh ...
NDUFS2 already contains a conserved protein kinase C phosphorylation site (soluble leptin receptor) at the C-terminus of the protein (Loeffen et al., 1998b); ...
[97]
Natural disease course and genotype-phenotype correlations in ...
May 30, 2012 · Mutations in one of the nuclear encoded structural or assembly genes of complex I have a dramatic effect on neurodevelopment and overall patient ...<|control11|><|separator|>
[98]
Lack of mitochondrial complex I assembly factor NDUFAF2 results in ...
Feb 28, 2024 · To date, only 14 patients with loss-of-function mutations in NDUFAF2 (including our 4 patients) have been reported.Missing: CRISPR | Show results with:CRISPR
[99]
CRISPR-Cas9 mediated knockout of NDUFS4 in human iPSCs
Here, we present a novel iPSC model of NDUFS4-related CI deficiency that displays a strong metabolic phenotype in the pluripotent state.