Abstract
The assembly of two deletion mutants of the Rieske iron-sulfur protein into the cytochrome bc 1 complex was investigated after import in vitro into mitochondria isolated from a strain of yeast, JPJ1, from which the iron-sulfur protein gene (RIP) had been deleted. The assembly process was investigated by immunoprecipitation of the labeled iron-sulfur protein or the two deletion mutants from detergent-solubilized mitochondria with specific antisera against either the iron-sulfur protein or the bc 1 complex (complex III) [Fu and Beattie (1991). J. Biol. Chem. 266, 16212–16218]. The deletion mutants lacking amino acid residues 55–66 or residues 161–180 were imported into mitochondria in vitro and processed to the mature form via an intermediate form. After import in vitro, the protein lacking residues 161–180 was not assembled into the complex, suggesting that the region of the iron-sulfur protein containing these residues may be involved in the assembly of the protein into the bc 1 complex; however, the protein lacking residues 55–66 was assembled in vitro into the bc 1 complex as effectively as the wild type iron-sulfur protein. Moreover, this mutant protein was present in the mitochondrial membrane fraction obtained from JPJ1 cells transformed with a single-copy plasmid containing the gene for this protein lacking residues 55–66. This deletion mutant protein was also assembled into the bc 1 complex in vivo, suggesting that the hydrophobic stretch of amino acids, residues 55–66, is not required for assembly of the iron-sulfur protein into the bc 1 complex; however, this association did not lead to enzymatic activity of the bc 1 complex, as the Rieske FeS cluster was not epr detectable in these mitochondria.
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Ramabadran, R.S., Japa, S. & Beattie, D.S. Assembly of Deletion Mutants of the Rieske Iron–Sulfur Protein into the Cytochrome bc 1 Complex of Yeast Mitochondria. J Bioenerg Biomembr 29, 45–54 (1997). https://doi.org/10.1023/A:1022459722020
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DOI: https://doi.org/10.1023/A:1022459722020