Effects of a missense exonic mutation in cytochrome b gene, observed on isolated mitochondrial complex III of Saccharomyces cerevisiae: consequence for the antimycin binding site
| Autor: | Paule Chevillotte-Brivet, Danielle Meunier-Lemesle, Nicole Forget, Gabrielle Salou |
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| Rok vydání: | 1987 |
| Předmět: |
Binding Sites
biology Cytochrome b Binding protein Cytochrome c Protein subunit Mutant Wild type Antimycin A General Medicine Exons Saccharomyces cerevisiae Cytochrome b Group Biochemistry Molecular biology Mitochondria chemistry.chemical_compound Electron Transport Complex III chemistry Genes Coenzyme Q – cytochrome c reductase Mutation biology.protein Heme |
| Zdroj: | Biochimie. 69(1) |
| ISSN: | 0300-9084 |
| Popis: | 1. 1. The mitochondrial complex III was isolated from a wild type strain of Saccharomyces cerevisiae PS409 and from two mutants, PS490 and PS493, carrying a missense mutation in the structural gene of cytochrome b (in exons B1 and B4 respectively). These mutants synthesize cytochrome b in variable proportions, but they are unable to grow on a respirable substrate. Strain PS493 does not bind antimycin, whereas strain PS490 contains less cytochromes b and c 1 but shows a strong binding to the inhibitor. 2. 2. The complex isolated from the wild type strain or mutant PS493 exhibited a specific cytochrome b and cytochrome c 1 heme content of approximately 8 and 4 nmol/mg of protein respectively. This content was about 3 and 2 nmol/mg with PS490, which leads to a molar stoichiometry of 1.3:1 for cytochromes b and c 1 , instead of an ‘ideal’ ratio of 2:1 expected with b–c 1 complex, and obtained with the two other strains. This implies that the association (or presence) of b and c 1 cytochromes is not pre-requisite for complex III assembly. 3. 3. The wild type complex III isolated from PS409 was found to have a high level of CoQ2H2 activity, using a synthetic coenzyme Q analog as substrate (440 s−1 mol of cytochrome c reduced/mol of cytochrome c 1 ). This activity is fully inhibited by antimycin. The complexes isolated from the two box mutants exhibited no such activity. 4. 4. Analysis of the subunit composition of the three isolated complexes on sodium dodecyl sulfate-gel electrophoresis showed that all the bands belonging to the b–c 1 complex were synthesized in both mutants as well as in the wild type strain. Some of them appeared to have slightly diminished, but no specific decrease of a band has been observed in mutant PS493 that does not bind antimycin, with respect to mutant PS490 which binds strongly to the inhibitor. It should be noted that the subunit of about 12–13 kDa, qualified as the antimycin binding protein, is equally present in the three complexes. 5. 5. The results suggest that the loss of antimycin binding in mutant PS493 might be due to conformational perturbations in the modified complex rather than to the disappearance or significant modification of some protein support. |
| Databáze: | OpenAIRE |
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