Heterologous complementation of a Rieske iron-sulfur protein-deficient Saccharomyces cerevisiae by the Rip1 gene of Schizosaccharomyces pombe.
A cDNA carrying the Rip1 gene, which encodes the Rieske iron-sulfur protein of Schizosaccharomyces pombe, has been cloned by complementing the respiratory deficiency of a Saccharomyces cerevisiae strain in which the endogenous copy of the RIP1 gene has been deleted. The deduced amino acid sequences of the S. pombe and S. cerevisiae iron-sulfur proteins are 50% identical, with the highest region of identity being in the C termini of the proteins, where the 2Fe:2S cluster is bound. When expressed in the S. cerevisiae deletion strain, the S. pombe iron-sulfur protein restores 25-30% of the ubiquinol-cytochrome c reductase activity. The kinetics of cytochrome c reduction, the effects of inhibitors which act at defined sites in the cytochrome bc1 complex, and the optical properties of cytochrome b in membranes from the S. cerevisiae deletion strain complemented with S. pombe iron-sulfur protein indicate that the S. pombe protein interacts with cytochrome b to restore an apparently normal ubiquinol oxidase site, but that interaction between the iron-sulfur protein and cytochrome c1 is partially impaired. This is the first heterologous replacement of an electron transfer protein in a respiratory enzyme complex in S. cerevisiae.
di Rago
J P
JP
Department of Biochemistry, Dartmouth Medical School, Hanover, New Hampshire 03755, USA.
Bruel
C
C
Graham
L A
LA
Slonimski
P
P
Trumpower
B L
BL
D000595:Amino Acid Sequence; D001483:Base Sequence; D003001:Cloning, Molecular; D014450:Electron Transport Complex III / Q000378:metabolism; D005800:Genes, Fungal; D005816:Genetic Complementation Test; D007506:Iron-Sulfur Proteins / Q000235:genetics*; D008969:Molecular Sequence Data; D010084:Oxidation-Reduction; D017433:Protein Structure, Secondary; D012441:Saccharomyces cerevisiae / Q000201:enzymology*; D012568:Schizosaccharomyces / Q000201:enzymology / Q000235:genetics*; D016415:Sequence Alignment; D017386:Sequence Homology, Amino Acid; D013057:Spectrum Analysis
D007506:Iron-Sulfur Proteins; C037989:Rieske iron-sulfur protein; D014450:Electron Transport Complex III