Molecular genetic studies of complex I inNeurospora crassa, Aspergillus niger andEscherichia coli

Weidner, Uwe; Nehls, Uwe; Schneider, Ralf; Fecke, Wolfgang; Leif, Hans; Schmiede, Andreas; Friedrich, Thorsten; Zensen, Ralf; Schulte, Ulrich; Ohnishi, Tomo̧ko; Weiss, Hanns

doi:10.1016/0005-2728(92)90218-q

Cited by 20 publications

(11 citation statements)

References 16 publications

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“…In addition, a second genetic locus (nuo) encoding a membrane-bound NADH dehydrogenase has been located at min 49 of the E. coli chromosome. Weidner et al have recently reported the cloning and partial DNA sequence of an E. coli gene which bears a high similarity to subunits of the mitochrondrial NADH dehydrogenase (complex I) (23).…”

Section: Resultsmentioning

confidence: 99%

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Demonstration of separate genetic loci encoding distinct membrane-bound respiratory NADH dehydrogenases in Escherichia coli

Calhoun

Gennis

1993

J Bacteriol

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The nature of the Escherichia coli membrane-bound NADH dehydrogenases and their role in the generation of the proton motive force has been controversial. One E. coli NADH:ubiquinone oxidoreductase has previously been purified to homogeneity, and its corresponding gene (ndh) has been isolated. However, two biochemically distinct E. coli NADH:ubiquinone oxidoreductase activities have been identified by others (K. Matsushita, T. Ohnishi, and H. R. Kaback, Biochemistry 26:7732-7737, 1987). An insertional mutation in the ndh gene has been introduced into the E. coli chromosome, and the resulting strain maintains membrane-bound NADH dehydrogenase activity, demonstrating that a second genetically distinct NADH dehydrogenase must be present. By standard genetic mapping techniques, the map position of a second locus (nuo) involved in the oxidation of NADH has been determined. The enzyme encoded by this locus probably translocates protons across the inner membrane, contributing to the proton motive force.The aerobic respiratory chain of Escherichia coli contains several dehydrogenases which catalyze the oxidation of various substrates. These dehydrogenases, including succinate dehydrogenase, D-lactate dehydrogenase, NADH dehydrogenase, and pyruvate oxidase, donate electrons to ubiquinone, which transfers the electrons to either of the two terminal oxidase complexes (1). The function of this electron transport chain is to generate a proton motive force across the membrane. This force, in turn, is responsible for a number of energy-dependent processes, such as ATP synthesis and active transport.The nature of the E. coli membrane-bound NADH dehydrogenases and their role in the generation of the proton motive force have been controversial. Young and Wallace isolated mutants deficient in membrane-bound NADH dehydrogenase activity by the inability of such mutants to utilize mannitol as the sole carbon source (28). These mutants were believed to contain single-locus mutations which were mapped to min 22 of the E. coli linkage map (2, 28). Subsequently, a gene encoding an E. coli NADH dehydrogenase (ndh) was cloned and sequenced (26,27). This membrane-associated NADH dehydrogenase was then purified from a strain harboring a recombinant plasmid (10). The purified enzyme (NDH) consists of a single polypeptide with a molecular mass of 47 kDa which contains flavin adenine dinucleotide but no iron (10, 11). It is highly active with ubiquinone-1 (Qi) as an electron acceptor (11). The data presented by Young and colleagues suggest that there are no iron-sulfur centers and no energy-coupling site associated with this NADH dehydrogenase.However, studies with inside-out membrane vesicles from E. coli strongly suggested that NADH oxidation is coupled to the generation of a proton motive force (14). In addition, Owen and Kaback (18)

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Section: Resultsmentioning

confidence: 99%

“…This second NADH dehydrogenase is of particular interest because of its similarities to its counterpart in mitochondria (23). The relative ease of manipulation of E. coli enzymes by molecular genetics makes NADH dhI an attractive model for the study of its mammalian counterpart.…”

Section: Resultsmentioning

confidence: 99%

Demonstration of separate genetic loci encoding distinct membrane-bound respiratory NADH dehydrogenases in Escherichia coli

Calhoun

Gennis

1993

J Bacteriol

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“…To establish a yeast genetic approach to complex I, our group has used the obligate aerobic yeast Yarrowia lipolytica [14], since this enzyme is absent in fermentative yeasts like Saccharomyces cerevisiae [15]. Further eucaryotic model organisms in complex I research include Bos taurus [16;17], fungi like Neurospora crassa [18], Podospora anserina [19] and Aspergillus niger [20], plants like Arabidopsis thaliana [21;22], Solanum tuberosum [23] and Oryza sativa [24], and many more. Chloroplasts and cyanobacteria possess a complex I like enzyme, which however has a different electron input module [25–27].…”

Section: Introductionmentioning

confidence: 99%

Architecture of complex I and its implications for electron transfer and proton pumping

Zickermann

Kerscher

Zwicker

et al. 2009

Biochimica et Biophysica Acta (BBA) - Bioenergetics

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Proton pumping NADH:ubiquinone oxidoreductase (complex I) is the largest and remains by far the least understood enzyme complex of the respiratory chain. It consists of a peripheral arm harbouring all known redox active prosthetic groups and a membrane arm with a yet unknown number of proton translocation sites. The ubiquinone reduction site close to iron-sulfur cluster N2 at the interface of the 49-kDa and PSST subunits has been mapped by extensive site directed mutagenesis. Independent lines of evidence identified electron transfer events during reduction of ubiquinone to be associated with the potential drop that generates the full driving force for proton translocation with a 4 H+/2e− stoichiometry. Electron microscopic analysis of immuno-labelled native enzyme and of a subcomplex lacking the electron input module indicated a distance of 35–60 Å of cluster N2 to the membrane surface. Resolution of the membrane arm into subcomplexes showed that even the distal part harbours subunits that are prime candidates to participate in proton translocation because they are homologous to sodium/proton antiporters and contain conserved charged residues in predicted transmembrane helices. The mechanism of redox linked proton translocation by complex I is largely unknown but has to include steps where energy is transmitted over extremely long distances. In this review we compile the available structural information on complex I and discuss implications for complex I function.

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“…The isolated mutant mitochondria respire as actively as the wild-type mitochondria with pyruvate/malate, NADH and succinate. Respiration of these substrates was inhibited by antimycin A or KCN, but was insensitive towards piericidin A. Electron paramagnetic resonance spectroscopy of the mutant mitochondria revealed only the signal of the iron-sulphur cluster N-2, but no signals arising from the other iron-sulphur clusters of complex I (see also Weidner et al, 1992). In summary, these data indicate that the mitochondria of mutant nu051 contain ubiquinol :cytochrome c oxidoreductase and cytochrome c oxidase in amounts similiar to the parental wildtype, contain about twice as much alternative NADH dehydrogenase, but do not contain a functional complex I.…”

Section: Some Properties Of the Mitochondria Isolated From The Mutantmentioning

confidence: 99%

The role of the proton‐pumping and alternative respiratory chain NADH:ubiquinone oxidoreductases in overflow catabolism of Aspergillus niger

Prömper

Schneider

Weiss

1993

European Journal of Biochemistry

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Mitochondria of fungi contain two respiratory chain enzymes concerned with the oxidation of matrix NADH. These are the proton-pumping NADH :ubiquinone oxidoreductase, also called complex I, which has a high affinity for NADH, and a non-proton-pumping NADH :ubiquinone oxidoreductase, called alternative NADH dehydrogenase, which has a low affinity for NADH. The role of these two enzymes in normal and overflow catabolism has been studied in Aspergillus niger. Three strains were investigated, the wild-type 732, the mutant nu051 that was generated from the wildtype by disrupting the gene of the (51-kDa) NADH-binding subunit of complex I and the citric acid over-producing strain B60 that looses complex I concomitantly with the onset of the over-production. Under standard growth conditions, respiratory energy transduction in the mutant nu051 was decreased by 40% compared to the parental wild-type and the strain B60. Respiratory electron transfer in the mutant nuo51, however, meets standard catabolic requirements. The intracellular levels of citric acid cycle intermediates in the mutant nu051 were the same as in the other two strains. Under growth conditions which lead to uncontrolled catabolic flux through glycolysis, a dramatic catabolic overflow occurred in the mutant nuo51. Intracellular levels of citric acid cycle intermediates increased to 20-fold normal levels. The strain B60, likewise lacking complex I under these conditions, excretes large amounts of citrate to moderate the intracellular catabolic overflow.The respiratory chain of filamentous fungi (and plants) contains, apart from the proton-pumping complexes NADH : ubiquinone oxidoreductase (complex I), ubiquino1 :cytochrome c oxidoreductase (complex 111) and cytochrome c oxidase (complex IVj, so-called alternative respiratory enzymes. Firstly, there are two additional NADH:ubiquinone oxidoreductases (alternative NADH dehydrogenases) ; one facing the matrix space of mitochondria and oxidizing internal NADH, the other facing the intermembrane space and oxidizing cytosolic NADH and/or NADPH. Secondly, there is a ubiquinol oxidase, also known as cyanide-resistant oxidase, transferring electrons from ubiquinol to oxygen directly. These alternative enzymes do not link electron transfer with proton translocation. They can bypass the proton-pumping complexes and, in terms of the energy transductional role of respiration, they are short circuits. Since these alternative respiratory enzymes have significantly lower affinities for

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Molecular genetic studies of complex I inNeurospora crassa, Aspergillus niger andEscherichia coli

Cited by 20 publications

References 16 publications

Demonstration of separate genetic loci encoding distinct membrane-bound respiratory NADH dehydrogenases in Escherichia coli

Demonstration of separate genetic loci encoding distinct membrane-bound respiratory NADH dehydrogenases in Escherichia coli

Architecture of complex I and its implications for electron transfer and proton pumping

The role of the proton‐pumping and alternative respiratory chain NADH:ubiquinone oxidoreductases in overflow catabolism of Aspergillus niger

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