Browse
Submit Data
Databases
API
Help

Dataset Information

0 Views

0 Connections

0 Citations

0 Reanalyses

0 Downloads

Omics score: 0

The 12.3 kDa subunit of complex I (respiratory-chain NADH dehydrogenase) from Neurospora crassa: cDNA cloning and chromosomal mapping of the gene.

ABSTRACT: The 12.3 kDa subunit of complex I (respiratory-chain NADH dehydrogenase) is a nuclear-coded protein of the hydrophobic fragment of the enzyme. We have isolated and sequenced a full-length cDNA clone coding for this polypeptide. The deduced protein is 104 amino acid residues long with a molecular mass of 12305 Da. This particular subunit of complex I lacks a cleavable mitochondrial targeting sequence. In agreement with its localization within complex I, we have found that this subunit behaves like an intrinsic membrane protein. Nevertheless, the deduced protein is rather hydrophilic, exhibiting no hydrophobic domain long enough to traverse a membrane in an alpha-helical conformation. The 12.3 kDa subunit shows a significant similarity to the hinge protein of complex III, suggesting that these two polypeptides may be involved in identical functions. This complex I subunit is coded for by a single gene. Applying restriction-fragment-length-polymorphism mapping, we located the gene on the right side of the centromere in linkage group I, linked to the lys-4 locus.

SUBMITTER: Videira A

PROVIDER: S-EPMC1132429 | biostudies-other | 1993 May

REPOSITORIES: biostudies-other

ACCESS DATA

Json Xml

Similar Datasets

Supramolecular organization of the respiratory chain in Neurospora crassa mitochondria.

Project description: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.

| S-EPMC2168242 | biostudies-literature

In organello assembly of respiratory-chain complex I: primary structure of the 14.8 kDa subunit of Neurospora crassa complex I.

Project description:A cDNA encoding the 14.8 kDa subunit of complex I from Neurospora crassa was cloned and sequenced. The deduced primary structure of this subunit reveals a predominantly hydrophilic protein containing no obvious membrane-spanning domain. In agreement with this characteristic, we have localized the 14.8 kDa subunit in the peripheral arm of the enzyme. The 14.8 kDa subunit was found to be conserved in mammalian complex I. The conservation of this subunit in such distantly related organisms suggests that the 14.8 kDa subunit is an important component of complex I. We have used an in organello system to study the biosynthetic pathway of this subunit. The 14.8 kDa polypeptide could be efficiently imported into isolated mitochondria. Furthermore, a fraction of the in-vitro-imported subunit was found to assemble in complex I. This is the first time that assembly in complex I of an in-vitro-synthesized subunit is demonstrated.

| S-EPMC1138054 | biostudies-other

Effects of disrupting the 21 kDa subunit of complex I from Neurospora crassa.

Project description:We have cloned and inactivated in vivo, by repeat-induced point mutations, the nuclear gene encoding a 21 kDa subunit of complex I from Neurospora crassa. Mitochondria from the nuo21 mutant lack this specific protein but retain other subunits of complex I in approximately normal amounts. In addition, this mutant is able to assemble an almost intact enzyme. The electron transfer activities from NADH to artificial acceptors of mitochondrial membranes from nuo21 differ from those of the wild-type strain, suggesting that the absence of the 21 kDa polypeptide results in conformational changes in complex I. Nevertheless, complex I of nuo21 is able to perform NADH:ubiquinone reductase activity, as judged by the observation that the respiration of mutant mitochondria is sensitive to inhibition by rotenone. We discuss these findings in relation to the involvement of complex I in mitochondrial diseases.

| S-EPMC1220496 | biostudies-other

Cryo-EM structure of <i>Neurospora crassa</i> respiratory complex IV.

Project description:In fungi, the mitochondrial respiratory chain complexes (complexes I-IV) are responsible for oxidative phosphorylation, as in higher eukaryotes. Cryo-EM was used to identify a 200 kDa membrane protein from Neurospora crassa in lipid nanodiscs as cytochrome c oxidase (complex IV) and its structure was determined at 5.5 Å resolution. The map closely resembles the cryo-EM structure of complex IV from Saccharomyces cerevisiae. Its ten subunits are conserved in S. cerevisiae and Bos taurus, but other transmembrane subunits are missing. The different structure of the Cox5a subunit is typical for fungal complex IV and may affect the interaction with complex III in a respiratory supercomplex. Additional density was found between the matrix domains of the Cox4 and Cox5a subunits that appears to be specific to N. crassa.

| S-EPMC6608615 | biostudies-literature

The internal alternative NADH dehydrogenase of Neurospora crassa mitochondria.

Project description:An open reading frame homologous with genes of non-proton-pumping NADH dehydrogenases was identified in the genome of Neurospora crassa. The 57 kDa NADH:ubiquinone oxidoreductase acts as internal (alternative) respiratory NADH dehydrogenase (NDI1) in the fungal mitochondria. The precursor polypeptide includes a pre-sequence of 31 amino acids, and the mature enzyme comprises one FAD molecule as a prosthetic group. It catalyses specifically the oxidation of NADH. Western blot analysis of fungal mitochondria fractionated with digitonin indicated that the protein is located at the inner face of the inner membrane of the organelle (internal enzyme). The corresponding gene was inactivated by the generation of repeat-induced point mutations. The respiratory activity of mitochondria from the resulting null-mutant ndi1 is almost fully inhibited by rotenone, an inhibitor of the proton-pumping complex I, when matrix-generated NADH is used as substrate. Although no effects of the NDI1 defect on vegetative growth and sexual differentiation were observed, the germination of both sexual and asexual ndi1 mutant spores is significantly delayed. Crosses between the ndi1 mutant strain and complex I-deficient mutants yielded no viable double mutants. Our data indicate: (i) that NDI1 represents the sole internal alternative NADH dehydrogenase of Neurospora mitochondria; (ii) that NDI1 and complex I are functionally complementary to each other; and (iii) that NDI1 is specially needed during spore germination.

| S-EPMC1223338 | biostudies-other

Mapping of lysine methylation and acetylation in core histones of Neurospora crassa.

Project description:Core histones are susceptible to a variety of post-translational modifications (PTMs), among which methylation and acetylation play critical roles in various chromatin-dependent processes. The nature and biological functions of these PTMs have been extensively studied in plants, animals, and yeasts. In contrast, the histone modifications in Neurospora crassa, a convenient model organism for multicellular eukaryotes, remained largely undefined. In this study, we used several mass spectrometric techniques, coupled with HPLC separation and multiple-protease digestion, to identify the methylation and acetylation sites in core histones isolated from Neurospora. Electron transfer dissociation (ETD) was employed to fragment the heavily modified long N-terminal peptides. In addition, accurate mass measurement of fragment ions allowed for unambiguous differentiation of acetylation from trimethylation. Many modification sites conserved in other organisms were identified in Neurospora. In addition, some unique modification sites in histone H2B, including N-terminal alpha methylation, methylation at K3, and acetylation at K19, K28, and K29, were observed. Our analysis provides a potentially comprehensive picture of methylation and acetylation of core histones in Neurospora, which should serve as a foundation for future studies of the function of histone PTMs in this model organism.

| S-EPMC2902163 | biostudies-literature

cDNA-derived amino acid sequence of the NADH-binding 51-kDa subunit of the bovine respiratory NADH dehydrogenase reveals striking similarities to a bacterial NAD(+)-reducing hydrogenase.

Project description:A lambda gt10 bovine brain and a lambda gt11 bovine heart cDNA library were screened with oligonucleotide probes corresponding to partial protein sequences directly determined from the isolated 51-kDa subunit of the bovine respiratory-chain NADH dehydrogenase. Clones were isolated that encode a protein of 464 amino acids containing all the 11 partial tryptic peptide sequences determined from the 51-kDa subunit. The size and amino acid composition of this protein agree with those determined for the purified 51-kDa subunit. Furthermore, this protein contains a putative NADH-binding domain, a possible FMN-binding site, and a putative binding site for an iron-sulfur cluster. The above evidence indicates that the cloned protein is the 51-kDa subunit or its precursor. A search for sequence similarity with proteins in the Protein Identification Resource data base has revealed that the 51-kDa subunit has 32% amino acid sequence identity with a major portion of the alpha subunit of the soluble NAD(+)-reducing hydrogenase from Alcaligenes eutrophus. In particular, there are three segments of high sequence similarity (70-88%) between the two proteins which correspond to the three ligand-binding sites.

| S-EPMC51631 | biostudies-other

Identification of a PRC2 Accessory Subunit Required for Subtelomeric H3K27 Methylation in Neurospora crassa.

Project description:Polycomb repressive complex 2 (PRC2) catalyzes methylation of histone H3 at lysine 27 (H3K27) in genomic regions of most eukaryotes and is critical for maintenance of the associated transcriptional repression. However, the mechanisms that shape the distribution of H3K27 methylation, such as recruitment of PRC2 to chromatin and/or stimulation of PRC2 activity, are unclear. Here, using a forward genetic approach in the model organism Neurospora crassa, we identified two alleles of a gene, NCU04278, encoding an unknown PRC2 accessory subunit (PAS). Loss of PAS resulted in losses of H3K27 methylation concentrated near the chromosome ends and derepression of a subset of associated subtelomeric genes. Immunoprecipitation followed by mass spectrometry confirmed reciprocal interactions between PAS and known PRC2 subunits, and sequence similarity searches demonstrated that PAS is not unique to N. crassa PAS homologs likely influence the distribution of H3K27 methylation and underlying gene repression in a variety of fungal lineages.

| S-EPMC7225565 | biostudies-literature

Secret lifestyles of Neurospora crassa.

Project description:Neurospora crassa has a long history as an excellent model for genetic, cellular, and biochemical research. Although this fungus is known as a saprotroph, it normally appears on burned vegetations or trees after forest fires. However, due to a lack of experimental evidence, the nature of its association with living plants remains enigmatic. Here we report that Scots pine (Pinus sylvestris) is a host plant for N. crassa. The endophytic lifestyle of N. crassa was found in its interaction with Scots pine. Moreover, the fungus can switch to a pathogenic state when its balanced interaction with the host is disrupted. Our data reveal previously unknown lifestyles of N. crassa, which are likely controlled by both environmental and host factors. Switching among the endophytic, pathogenic, and saprotrophic lifestyles confers upon fungi phenotypic plasticity in adapting to changing environments and drives the evolution of fungi and associated plants.

| S-EPMC4038807 | biostudies-literature

NADH:ubiquinone oxidoreductase from bovine mitochondria. cDNA sequence of a 19 kDa cysteine-rich subunit.

Project description:The sequence of a 19 kDa subunit of NADH:ubiquinone oxidoreductase (complex I) from bovine heart mitochondria has been determined by a new strategy based on the polymerase chain reaction. The subunits of the enzyme were resolved in a polyacrylamide gel by two-dimensional isoelectric focusing and electrophoresis under denaturing conditions, transferred to a poly(vinylidene difluoride) membrane, and the N-terminal sequence was determined on the stained 19 kDa protein up to residue 27. This information was used to design two mixed oligonucleotide primers and a mixed oligonucleotide probe. With total bovine heart cDNA as template, overlapping cDNAs extending to sequences corresponding to both the 5' and 3' extremities of the mRNA coding for the 19 kDa subunit were synthesized in three polymerase chain reactions. These cDNAs were cloned and sequenced and encode a 171-amino-acid mature protein preceded by a methionine residue. The mature protein contains eight cysteine residues spaced at regular intervals through the protein, but the cysteine-rich motifs that are often associated with tetranuclear or binuclear centres in other proteins are not present. However, all eight cysteine residues are strictly conserved in a related protein from Neurospora crassa, suggesting that they have structural and/or functional significance in complex I.

| S-EPMC1151184 | biostudies-other

OmicsDI is part of the ELIXIR infrastructure

OmicsDI is an Elixir interoperability service. Learn more ›

Tweets

OmicsDI Databases

PRIDE
PeptideAtlas
MassIVE
JPOST Repository
Physiome Model Repository

EGA
EVA
ENA
LINCS
PAXDB
Cell Collective

MetaboLights
Metabolomics Workbench
MetabolomeExpress
GNPS
BioModels
FAIRDOMHub

ArrayExpress
dbGaP
ExpressionAtlas
GEO
NODE

Information

Databases
Help
API
Contact us
Code on GitHub
Terms of use
Submit Data