We have analyzed the structure/function relationships of the yeast mitochondrial cytochrome b with a new methodology based upon the isolation of pseudo-wild type revertants from well-characterized cytochrome b respiratory deficient mutants. Our goal was to determine how cytochrome b function could be restored in such mutants, at least to some degree, by suppressor mutations within the protein. True wild type revertants were differentiated from pseudo-wild type revertants by the use of a simple and rapid screening technique based upon oligonucleotide hybridization. This can easily be used to analyze a large number of revertants. The suppressor mutations responsible for the restoration of respiratory competence were identified by sequencing the revertant's cytochrome b mRNA in crude mitochondrial RNA preparations. Using this new method we have analyzed 210 independent revertants. We report here nine novel cytochrome b structures conferring a variety of respiratory sufficient phenotypes, obtained from five respiratory deficient mutations affecting a short region of the protein (positions 131-138 of the polypeptide chain), presumably belonging to the ubiquinol oxidizing center of the bc1 complex.

• PMID: 2154475
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Comparative Study | Journal Article | Research Support, Non-U.S. Gov't

Authors

di Rago JP, Netter P, Slonimski PP

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