Subunit arrangement in beef heart complex III

Energy transduction in mitochondria involves five oligomeric complexes embedded within the inner membrane. They are composed of catalytic and noncatalytic subunits, the role of these latter proteins often being difficult to assign. One of these complexes, the bc1 complex, is composed of three catalytic subunits including cytochrome b and seven or eight noncatalytic subunits. Recently, several mutations in the human cytochrome b gene have been linked to various diseases. We have studied in detail the effects of a cardiomyopathy generating mutation G252D in yeast. This mutation disturbs the biogenesis of the bc1 complex at 36°C and decreases the steady-state level of the noncatalytic subunit Qcr9p. In addition, the G252D mutation and the deletion of QCR9 show synergetic defects that can be partially bypassed by suppressor mutations at position 252 and by a new cytochrome b mutation, P174T. Altogether, our results suggest that the supernumerary subunit Qcr9p enhances or stabilizes the interactions between the catalytic subunits, this role being essential at high temperature.Numerous cellular functions are performed by oligomeric complexes composed of catalytic and noncatalytic subunits and the role of these latter proteins is often difficult to assign. Energy transduction in mitochondria involves five oligomeric complexes that are embedded within the inner membrane. One of these complexes, the bc1 complex is composed of three catalytic and seven or eight noncatalytic subunits (for review, see Ref. 1). The three catalytic subunits, cytochrome b, cytochrome c1, and the Rieske iron-sulfur protein (Rieske) are conserved in the bacterial equivalents of this complex. However, several noncatalytic subunits are present in the mitochondrial bc1 complexes and are often referred to as supernumerary subunits although some are required for the enzymatic activity. In Saccharomyces cerevisiae the bc1 complex contains seven supernumerary subunits that are conserved in the mammalian enzyme. The mammalian bc1 complex presents an additional subunit that has no equivalent in yeast. The mitochondrial bc1 complexes from bovine, chicken, and yeast have been crystallized (2-5) and an analysis of these structures reveals that the general shape, size, and topology of the complexes are similar in the three organisms.

Section: Discussionmentioning

confidence: 69%

A Pathogenic Cytochrome b Mutation Reveals New Interactions between Subunits of the Mitochondrial bc1Complex

Saint-Georges¹,

Bonnefoy²,

Rago³

et al. 2002

“…Apparent molecular masses were calculated based on the reported molecular masses of bovine cytochrome c oxidase (24). Immunoblotting was carried out as in Gonzá lez-Halphen et al (25). Antibodies against yeast COX III were obtained from Molecular Probes (Eugene, Oregon).…”

Section: Methodsmentioning

confidence: 99%

Unusual Location of a Mitochondrial Gene

Pérez-Martı́nez

Vázquez-Acevedo

Tolkunova

et al. 2000

The algae of the family Chlamydomonadaceae lack the gene cox3 that encodes subunit III of cytochrome c oxidase in their mitochondrial genomes. This observation has raised the question of whether this subunit is present in cytochrome c oxidase or whether the corresponding gene is located in the nucleus. Cytochrome c oxidase was isolated from the colorless chlamydomonad Polytomella spp., and the existence of subunit III was established by immunoblotting analysis with an antibody directed against Saccharomyces cerevisiae subunit III. Based partly upon the N-terminal sequence of this subunit, oligodeoxynucleotides were designed and used for polymerase chain reaction amplification, and the resulting product was used to screen a cDNA library of Chlamydomonas reinhardtii. The complete sequences of the cox3 cDNAs from Polytomella spp. and C. reinhardtii are reported. Evidence is provided that the genes for cox3 are encoded by nuclear DNA, and the predicted polypeptides exhibit diminished physical constraints for import as compared with mitochondrial-DNA encoded homologs. This indicates that transfer of this gene to the nucleus occurred before Polytomella diverged from the photosynthetic Chlamydomonas lineage and that this transfer may have occurred in all chlamydomonad algae.

“…Antibodies were raised in rabbits injected with purified yeast GDH1-encoded NADP-GDH and partially purified by ammonium sulfate precipitation according to the method of Gonzá lez-Halphen et al (26).…”

Section: Preparation Of Anti-nadp-gdh Antibodiesmentioning

confidence: 99%

NADP-Glutamate Dehydrogenase Isoenzymes of Saccharomyces cerevisiae

DeLuna¹,

Avendaño

Riego

et al. 2001

141

In the yeast Saccharomyces cerevisiae, two NADP؉ -dependent glutamate dehydrogenases (NADP-GDHs) encoded by GDH1 and GDH3 catalyze the synthesis of glutamate from ammonium and ␣-ketoglutarate. The GDH2-encoded NAD ؉ -dependent glutamate dehydrogenase degrades glutamate producing ammonium and ␣-ketoglutarate. Until very recently, it was considered that only one biosynthetic NADP-GDH was present in S. cerevisiae. This fact hindered understanding the physiological role of each isoenzyme and the mechanisms involved in ␣-ketoglutarate channeling for glutamate biosynthesis. In this study, we purified and characterized the GDH1-and GDH3-encoded NADP-GDHs; they showed different allosteric properties and rates of ␣-ketoglutarate utilization. Analysis of the relative levels of these proteins revealed that the expression of GDH1 and GDH3 is differentially regulated and depends on the nature of the carbon source. Moreover, the physiological study of mutants lacking or overexpressing GDH1 or GDH3 suggested that these genes play nonredundant physiological roles. Our results indicate that the coordinated regulation of GDH1-, GDH3-, and GDH2-encoded enzymes results in glutamate biosynthesis and balanced utilization of ␣-ketoglutarate under fermentative and respiratory conditions. The possible relevance of the duplicated NADP-GDH pathway in the adaptation to facultative metabolism is discussed.