scholarly journals The Campylobacter jejuni NADH:Ubiquinone Oxidoreductase (Complex I) Utilizes Flavodoxin Rather than NADH

2007 ◽  
Vol 190 (3) ◽  
pp. 915-925 ◽  
Author(s):  
Dilan R. Weerakoon ◽  
Jonathan W. Olson
Keyword(s):  
Campylobacter Jejuni ◽  
Complex I ◽  
Nadh Dehydrogenase ◽  
Respiratory Activity ◽  
Essential Gene ◽  
Electron Flow ◽  
Nadh:Ubiquinone Oxidoreductase ◽  
The Novel ◽  
Wild Type ◽  
Transport Chain

ABSTRACT Campylobacter jejuni encodes 12 of the 14 subunits that make up the respiratory enzyme NADH:ubiquinone oxidoreductase (also called complex I). The two nuo genes not present in C. jejuni encode the NADH dehydrogenase, and in their place in the operon are the novel genes designated Cj1575c and Cj1574c. A series of mutants was generated in which each of the 12 nuo genes (homologues to known complex I subunits) was disrupted or deleted. Each of the nuo mutants will not grow in amino acid-based medium unless supplemented with an alternative respiratory substrate such as formate. Unlike the nuo genes, Cj1574c is an essential gene and could not be disrupted unless an intact copy of the gene was provided at an unrelated site on the chromosome. A nuo deletion mutant can efficiently respire formate but is deficient in α-ketoglutarate respiratory activity compared to the wild type. In C. jejuni, α-ketoglutarate respiration is mediated by the enzyme 2-oxoglutarate:acceptor oxidoreductase; mutagenesis of this enzyme abolishes α-ketoglutarate-dependent O2 uptake and fails to reduce the electron transport chain. The electron acceptor for 2-oxoglutarate:acceptor oxidoreductase was determined to be flavodoxin, which was also determined to be an essential protein in C. jejuni. A model is presented in which CJ1574 mediates electron flow into the respiratory transport chain from reduced flavodoxin and through complex I.

scholarly journals The mitochondrial AAA protease FTSH3 regulates Complex I abundance by promoting its disassembly

2021 ◽  
Author(s):  
Aneta Ivanova ◽  
Abi S Ghifari ◽  
Oliver Berkowitz ◽  
James Whelan ◽  
Monika W Murcha
Keyword(s):  
Complex I ◽  
Nadh Dehydrogenase ◽  
Slow Growth ◽  
Nadh:Ubiquinone Oxidoreductase ◽  
Temperature Sensitive ◽  
Wild Type ◽  
Forward Genetic Screen ◽  
Slow Growth Rate ◽  
Partial Restoration ◽  
Screen Approach

Abstract ATP is generated in mitochondria by oxidative phosphorylation. Complex I (NADH:ubiquinone oxidoreductase or NADH dehydrogenase) is the first multisubunit protein complex of this pathway, oxidising NADH and transferring electrons to the ubiquinone pool. Typically Complex I mutants display a slow growth rate compared to wild-type plants. Here, using a forward genetic screen approach for restored growth of a Complex I mutant, we have identified the mitochondrial ATP dependent metalloprotease, Filamentous Temperature Sensitive H 3 (FTSH3), as a factor that is required for the disassembly of Complex I. An ethyl methanesulfonate-induced mutation in FTSH3, named rmb1 (restoration of mitochondrial biogenesis 1), restored Complex I abundance and plant growth. Complementation could be achieved with FTSH3 lacking proteolytic activity, suggesting the unfoldase function of FTSH3 has a role in Complex I disassembly. The introduction of the rmb1 to an additional, independent, and extensively characterised Complex I mutant, ndufs4, resulted in similar increases to Complex I abundance and a partial restoration of growth. These results show that disassembly or degradation of Complex I plays a role in determining its steady-state abundance and thus turnover may vary under different conditions.

scholarly journals Role of Campylobacter jejuni Respiratory Oxidases and Reductases in Host Colonization

2008 ◽  
Vol 74 (5) ◽  
pp. 1367-1375 ◽  
Author(s):  
Rebecca A. Weingarten ◽  
Jesse L. Grimes ◽  
Jonathan W. Olson
Keyword(s):  
Campylobacter Jejuni ◽  
Anaerobic Respiration ◽  
Wild Type ◽  
Respiratory Enzymes ◽  
Host Colonization ◽  
Enzymatic Function ◽  
Food Borne ◽  
Transport Chain ◽  
Respiratory Oxidases

ABSTRACT Campylobacter jejuni is the leading cause of human food-borne bacterial gastroenteritis. The C. jejuni genome sequence predicts a branched electron transport chain capable of utilizing multiple electron acceptors. Mutants were constructed by disrupting the coding regions of the respiratory enzymes nitrate reductase (napA::Cm), nitrite reductase (nrfA::Cm), dimethyl sulfoxide, and trimethylamine N-oxide reductase (termed Cj0264::Cm) and the two terminal oxidases, a cyanide-insensitive oxidase (cydA::Cm) and cbb3-type oxidase (ccoN::Cm). Each strain was characterized for the loss of the associated enzymatic function in vitro. The strains were then inoculated into 1-week-old chicks, and the cecal contents were assayed for the presence of C. jejuni 2 weeks postinoculation. cydA::Cm and Cj0264c::Cm strains colonized as well as the wild type; napA::Cm and nrfA::Cm strains colonized at levels significantly lower than the wild type. The ccoN::Cm strain was unable to colonize the chicken; no colonies were recovered at the end of the experiment. While there appears to be a role for anaerobic respiration in host colonization, oxygen is the most important respiratory acceptor for C. jejuni in the chicken cecum.

scholarly journals Oxidation and reduction of pyridine nucleotides in alamethicin-permeabilized plant mitochondria

2004 ◽  
Vol 380 (1) ◽  
pp. 193-202 ◽  
Author(s):  
Fredrik I. JOHANSSON ◽  
Agnieszka M. MICHALECKA ◽  
Ian M. MØLLER ◽  
Allan G. RASMUSSON
Keyword(s):  
Electron Transport ◽  
Complex I ◽  
Electron Transport Chain ◽  
Nadh Dehydrogenase ◽  
Plant Mitochondria ◽  
Inner Mitochondrial Membrane ◽  
Intact Cells ◽  
Transport Chain ◽  
Invasive Method

The inner mitochondrial membrane is selectively permeable, which limits the transport of solutes and metabolites across the membrane. This constitutes a problem when intramitochondrial enzymes are studied. The channel-forming antibiotic AlaM (alamethicin) was used as a potentially less invasive method to permeabilize mitochondria and study the highly branched electron-transport chain in potato tuber (Solanum tuberosum) and pea leaf (Pisum sativum) mitochondria. We show that AlaM permeabilized the inner membrane of plant mitochondria to NAD(P)H, allowing the quantification of internal NAD(P)H dehydrogenases as well as matrix enzymes in situ. AlaM was found to inhibit the electron-transport chain at the external Ca2+-dependent rotenone-insensitive NADH dehydrogenase and around complexes III and IV. Nevertheless, under optimal conditions, especially complex I-mediated NADH oxidation in AlaM-treated mitochondria was much higher than what has been previously measured by other techniques. Our results also show a difference in substrate specificities for complex I in mitochondria as compared with inside-out submitochondrial particles. AlaM facilitated the passage of cofactors to and from the mitochondrial matrix and allowed the determination of NAD+ requirements of malate oxidation in situ. In summary, we conclude that AlaM provides the best method for quantifying NADH dehydrogenase activities and that AlaM will prove to be an important method to study enzymes under conditions that resemble their native environment not only in plant mitochondria but also in other membrane-enclosed compartments, such as intact cells, chloroplasts and peroxisomes.

scholarly journals Tracking Electron Uptake from a Cathode into Shewanella Cells: Implications for Energy Acquisition from Solid-Substrate Electron Donors

mBio ◽  
2018 ◽  
Vol 9 (1) ◽  
Author(s):  
Annette R. Rowe ◽  
Pournami Rajeev ◽  
Abhiney Jain ◽  
Sahand Pirbadian ◽  
Akihiro Okamoto ◽  
...  
Keyword(s):  
Electron Transfer ◽  
Electron Transport ◽  
Complex I ◽  
Electron Transport Chain ◽  
Electron Flow ◽  
Extracellular Electron Transfer ◽  
Terminal Electron Acceptor ◽  
Cellular Energy ◽  
Transport Chain ◽  
Energy Acquisition

ABSTRACTWhile typically investigated as a microorganism capable of extracellular electron transfer to minerals or anodes,Shewanella oneidensisMR-1 can also facilitate electron flow from a cathode to terminal electron acceptors, such as fumarate or oxygen, thereby providing a model system for a process that has significant environmental and technological implications. This work demonstrates that cathodic electrons enter the electron transport chain ofS. oneidensiswhen oxygen is used as the terminal electron acceptor. The effect of electron transport chain inhibitors suggested that a proton gradient is generated during cathode oxidation, consistent with the higher cellular ATP levels measured in cathode-respiring cells than in controls. Cathode oxidation also correlated with an increase in the cellular redox (NADH/FMNH2) pool determined with a bioluminescence assay, a proton uncoupler, and a mutant of proton-pumping NADH oxidase complex I. This work suggested that the generation of NADH/FMNH2under cathodic conditions was linked to reverse electron flow mediated by complex I. A decrease in cathodic electron uptake was observed in various mutant strains, including those lacking the extracellular electron transfer components necessary for anodic-current generation. While no cell growth was observed under these conditions, here we show that cathode oxidation is linked to cellular energy acquisition, resulting in a quantifiable reduction in the cellular decay rate. This work highlights a potential mechanism for cell survival and/or persistence on cathodes, which might extend to environments where growth and division are severely limited.IMPORTANCEThe majority of our knowledge of the physiology of extracellular electron transfer derives from studies of electrons moving to the exterior of the cell. The physiological mechanisms and/or consequences of the reverse processes are largely uncharacterized. This report demonstrates that when coupled to oxygen reduction, electrode oxidation can result in cellular energy acquisition. This respiratory process has potentially important implications for how microorganisms persist in energy-limited environments, such as reduced sediments under changing redox conditions. From an applied perspective, this work has important implications for microbially catalyzed processes on electrodes, particularly with regard to understanding models of cellular conversion of electrons from cathodes to microbially synthesized products.

scholarly journals Supramolecular Organization of the Respiratory Chain in Neurospora crassa Mitochondria

2007 ◽  
Vol 6 (12) ◽  
pp. 2391-2405 ◽  
Author(s):  
Isabel Marques ◽  
Norbert A. Dencher ◽  
Arnaldo Videira ◽  
Frank Krause
Keyword(s):  
Neurospora Crassa ◽  
Respiratory Chain ◽  
Complex I ◽  
Polyacrylamide Gel Electrophoresis ◽  
Nadh:Ubiquinone Oxidoreductase ◽  
Deletion Mutants ◽  
Supramolecular Organization ◽  
Wild Type ◽  
Respiratory Supercomplexes ◽  
Native Polyacrylamide Gel Electrophoresis

ABSTRACT The existence of specific respiratory supercomplexes in mitochondria of most organisms has gained much momentum. However, its functional significance is still poorly understood. The availability of many deletion mutants in complex I (NADH:ubiquinone oxidoreductase) of Neurospora crassa, distinctly affected in the assembly process, offers unique opportunities to analyze the biogenesis of respiratory supercomplexes. Herein, we describe the role of complex I in assembly of respiratory complexes and supercomplexes as suggested by blue and colorless native polyacrylamide gel electrophoresis and mass spectrometry analyses of mildly solubilized mitochondria from the wild type and eight deletion mutants. As an important refinement of the fungal respirasome model, we found that the standard respiratory chain of N. crassa comprises putative complex I dimers in addition to I-III-IV and III-IV supercomplexes. Three Neurospora mutants able to assemble a complete complex I, lacking only the disrupted subunit, have respiratory supercomplexes, in particular I-III-IV supercomplexes and complex I dimers, like the wild-type strain. Furthermore, we were able to detect the I-III-IV supercomplexes in the nuo51 mutant with no overall enzymatic activity, representing the first example of inactive respirasomes. In addition, III-IV supercomplexes were also present in strains lacking an assembled complex I, namely, in four membrane arm subunit mutants as well as in the peripheral arm nuo30.4 mutant. In membrane arm mutants, high-molecular-mass species of the 30.4-kDa peripheral arm subunit comigrating with III-IV supercomplexes and/or the prohibitin complex were detected. The data presented herein suggest that the biogenesis of complex I is linked with its assembly into supercomplexes.

scholarly journals Structural organization of mitochondrial human complex I: role of the ND4 and ND5 mitochondria-encoded subunits and interaction with prohibitin

2004 ◽  
Vol 383 (3) ◽  
pp. 491-499 ◽  
Author(s):  
Ingrid BOURGES ◽  
Claire RAMUS ◽  
Bénédicte MOUSSON de CAMARET ◽  
Réjane BEUGNOT ◽  
Claire REMACLE ◽  
...  
Keyword(s):  
Complex I ◽  
Nadh Dehydrogenase ◽  
Nadh:Ubiquinone Oxidoreductase ◽  
Membrane Association ◽  
Oxidoreductase Activity ◽  
Mitochondrial Inner Membrane ◽  
The Matrix ◽  
Complex I Assembly ◽  
Human Complex

Mitochondria-encoded ND (NADH dehydrogenase) subunits, as components of the hydrophobic part of complex I, are essential for NADH:ubiquinone oxidoreductase activity. Mutations or lack of expression of these subunits have significant pathogenic consequences in humans. However, the way these events affect complex I assembly is poorly documented. To understand the effects of particular mutations in ND subunits on complex I assembly, we studied four human cell lines: ND4 non-expressing cells, ND5 non-expressing cells, and rho° cells that do not express any ND subunits, in comparison with normal complex I control cells. In control cells, all the seven analysed nuclear-encoded complex I subunits were found to be attached to the mitochondrial inner membrane, except for the 24 kDa subunit, which was nearly equally partitioned between the membranes and the matrix. Absence of a single ND subunit, or even all the seven ND subunits, caused no major changes in the nuclear-encoded complex I subunit content of mitochondria. However, in cells lacking ND4 or ND5, very low amounts of 24 kDa subunit were found associated with the membranes, whereas most of the other nuclear-encoded subunits remained attached. In contrast, membrane association of most of the nuclear subunits was significantly reduced in the absence of all seven ND proteins. Immunopurification detected several subcomplexes. One of these, containing the 23, 30 and 49 kDa subunits, also contained prohibitin. This is the first description of prohibitin interaction with complex I subunits and suggests that this protein might play a role in the assembly or degradation of mitochondrial complex I.

scholarly journals Evolution of an assembly factor-based subunit contributed to a novel NDH-PSI supercomplex formation in chloroplasts

2021 ◽  
Vol 12 (1) ◽  
Author(s):  
Yoshinobu Kato ◽  
Masaki Odahara ◽  
Toshiharu Shikanai
Keyword(s):  
Nadh Dehydrogenase ◽  
Common Ancestor ◽  
Electron Flow ◽  
Chain Complex ◽  
Photosynthetic Electron Transport ◽  
Mitochondrial Complex ◽  
Contact Site ◽  
Tandem Gene Duplication ◽  
Transport Chain ◽  
Assembly Factor

AbstractChloroplast NADH dehydrogenase-like (NDH) complex is structurally related to mitochondrial Complex I and forms a supercomplex with two copies of Photosystem I (the NDH-PSI supercomplex) via linker proteins Lhca5 and Lhca6. The latter was acquired relatively recently in a common ancestor of angiosperms. Here we show that NDH-dependent Cyclic Electron Flow 5 (NDF5) is an NDH assembly factor in Arabidopsis. NDF5 initiates the assembly of NDH subunits (PnsB2 and PnsB3) and Lhca6, suggesting that they form a contact site with Lhca6. Our analysis of the NDF5 ortholog in Physcomitrella and angiosperm genomes reveals the subunit PnsB2 to be newly acquired via tandem gene duplication of NDF5 at some point in the evolution of angiosperms. Another Lhca6 contact subunit, PnsB3, has evolved from a protein unrelated to NDH. The structure of the largest photosynthetic electron transport chain complex has become more complicated by acquiring novel subunits and supercomplex formation with PSI.

scholarly journals Inactivation of mitochondrial Complex I stimulates chloroplast ATPase in Physcomitrella (Physcomitrium patens)

2020 ◽  
Author(s):  
Marco Mellon ◽  
Mattia Storti ◽  
Antoni Mateu Vera Vives ◽  
David M. Kramer ◽  
Alessandro Alboresi ◽  
...  
Keyword(s):  
Complex I ◽  
Respiratory Activity ◽  
Photosynthetic Apparatus ◽  
Mitochondrial Complex ◽  
Photosynthetic Organisms ◽  
Photosystems I And Ii ◽  
Transport Chain ◽  
Ultimate Source ◽  
Photosynthetic Control ◽  
Atp Supply

AbstractWhile light is the ultimate source of energy for photosynthetic organisms, mitochondrial respiration is still fundamental for supporting metabolism demand during the night or in heterotrophic tissues. Respiration is also important for the metabolism of photosynthetically active cells, acting as a sink for excess reduced molecules and source of substrates for anabolic pathways. In this work, we isolated Physcomitrella (Physcomitrium patens) plants with altered respiration by inactivating Complex I of the mitochondrial electron transport chain by independent targeting of two essential subunits. Results show that the inactivation of Complex I causes a strong growth impairment even in fully autotrophic conditions in tissues where all cells are photosynthetically active. Complex I mutants show major alterations in the stoichiometry of respiratory complexes while the composition of photosynthetic apparatus was substantially unaffected. Complex I mutants showed altered photosynthesis with higher yields of both Photosystems I and II. These are the consequence of a higher chloroplast ATPase activity that also caused a smaller ΔpH formation across thylakoid membranes as well as decreased photosynthetic control on cytochrome b6f, possibly to compensate for a deficit in ATP supply relative to demand in Complex I mutants. These results demonstrate that alteration of respiratory activity directly impacts photosynthesis in P. patens and that metabolic interaction between organelles is essential in their ability to use light energy for growth.

scholarly journals Novel mitochondrial complex I-inhibiting peptides restrain NADH dehydrogenase activity

2019 ◽  
Vol 9 (1) ◽  
Author(s):  
Yao-Peng Xue ◽  
Mou-Chieh Kao ◽  
Chung-Yu Lan
Keyword(s):  
Antimicrobial Peptides ◽  
Complex I ◽  
Fungal Pathogens ◽  
Nadh Dehydrogenase ◽  
Mitochondrial Complex I ◽  
Mitochondrial Complex ◽  
Human Fungal Pathogen ◽  
New Class ◽  
Transport Chain ◽  
Nadh Dehydrogenases

Abstract The emergence of drug-resistant fungal pathogens is becoming increasingly serious due to overuse of antifungals. Antimicrobial peptides have potent activity against a broad spectrum of pathogens, including fungi, and are considered a potential new class of antifungals. In this study, we examined the activities of the newly designed peptides P-113Du and P-113Tri, together with their parental peptide P-113, against the human fungal pathogen Candida albicans. The results showed that these peptides inhibit mitochondrial complex I, specifically NADH dehydrogenase, of the electron transport chain. Moreover, P-113Du and P-113Tri also block alternative NADH dehydrogenases. Currently, most inhibitors of the mitochondrial complex I are small molecules or artificially-designed antibodies. Here, we demonstrated novel functions of antimicrobial peptides in inhibiting the mitochondrial complex I of C. albicans, providing insight in the development of new antifungal agents.

scholarly journals Assembly of Mitochondrial Complex I Requires the Low-Complexity Protein AMC1 in Chlamydomonas reinhardtii

Genetics ◽  
2020 ◽  
Vol 214 (4) ◽  
pp. 895-911 ◽  
Author(s):  
Nitya Subrahmanian ◽  
Andrew David Castonguay ◽  
Claire Remacle ◽  
Patrice Paul Hamel
Keyword(s):  
Chlamydomonas Reinhardtii ◽  
Complex I ◽  
Mitochondrial Gene ◽  
Low Complexity ◽  
Wild Type ◽  
Reporter Protein ◽  
Transport Chain ◽  
Type Gene ◽  
Complex I Assembly ◽  
Assembly Defect

Complex I is the first enzyme involved in the mitochondrial electron transport chain. With >40 subunits of dual genetic origin, the biogenesis of complex I is highly intricate and poorly understood. We used Chlamydomonas reinhardtii as a model system to reveal factors involved in complex I biogenesis. Two insertional mutants, displaying a complex I assembly defect characterized by the accumulation of a 700 kDa subcomplex, were analyzed. Genetic analyses showed these mutations were allelic and mapped to the gene AMC1 (Cre16.g688900) encoding a low-complexity protein of unknown function. The complex I assembly and activity in the mutant was restored by complementation with the wild-type gene, confirming AMC1 is required for complex I biogenesis. The N terminus of AMC1 targets a reporter protein to yeast mitochondria, implying that AMC1 resides and functions in the Chlamydomonas mitochondria. Accordingly, in both mutants, loss of AMC1 function results in decreased abundance of the mitochondrial nd4 transcript, which encodes the ND4 membrane subunit of complex I. Loss of ND4 in a mitochondrial nd4 mutant is characterized by a membrane arm assembly defect, similar to that exhibited by loss of AMC1. These results suggest AMC1 is required for the production of mitochondrially-encoded complex I subunits, specifically ND4. We discuss the possible modes of action of AMC1 in mitochondrial gene expression and complex I biogenesis.

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