Genetic analysis of yeast iso-1-cytochrome c structural requirements: suppression of Gly6 replacements by an Asn52----Ile replacement.
Gly6 (vertebrate numbering system) is an evolutionarily invariant amino acid located in an electron-dense region of cytochrome c. Serine, cysteine, and aspartic acid replacements of Gly6 abolished yeast iso-1-cytochrome c function, presumably by destabilizing the mature forms of the altered proteins (1). Here we report that genetic reversion analysis of these mutants has uncovered a single base-pair substitution, encoding an Asn52----Ile replacement, that suppresses all three position 6 defects, as well as a Gly6....Gly29----Ser6....Ser29 double replacement. In each case the suppressor restored at least partial function to the altered iso-1-cytochromes c, with the Sera6....Ile52 protein being nearly indistinguishable from the normal protein. The suppressor also affected otherwise normal iso-1-cytochrome c, enhancing the in vivo amount of the protein by about 20%. While this work was in progress, Das et al. (1989, Proc. Natl. Acad. Sci. USA 86, 496-499) uncovered Ile52 as a suppressor of single Gly29 and His33 replacements in iso-1-cytochrome c. The ability of Ile52 to suppress amino acid replacements at three different sites, and its effect in isolation from the primary mutations, defines Ile52 as a global suppressor of specific iso-1-cytochrome c structural defects. These data suggest that position 52 plays a critical role in the folding and/or stability of iso-1-cytochrome c.
Berroteran
R W
RW
Department of Biochemistry and Molecular Biology, Louisiana State University Medical Center, Shreveport 71130.
Hampsey
M
M
D001483:Base Sequence; D005075:Biological Evolution; D003001:Cloning, Molecular; D003574:Cytochrome c Group / Q000737:chemistry / Q000235:genetics*; D045304:Cytochromes c; D004252:DNA Mutational Analysis; D004271:DNA, Fungal / Q000235:genetics; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D011487:Protein Conformation; D012441:Saccharomyces cerevisiae / Q000201:enzymology* / Q000235:genetics; D029701:Saccharomyces cerevisiae Proteins; D013489:Suppression, Genetic
C014059:CYC1 protein, S cerevisiae; D003574:Cytochrome c Group; D004271:DNA, Fungal; D029701:Saccharomyces cerevisiae Proteins; D045304:Cytochromes c