Dramatic thermostabilization of yeast iso-1-cytochrome c by an asparagine----isoleucine replacement at position 57.

Abstract

Two Saccharomyces cerevisiae yeast mutants, cyc1-73 and cyc1-190, contain nonfunctional and presumably unstable forms of iso-1-cytochrome c due to Gly-34.fwdarw. Ser and His-38.fwdarw. Pro replacements, respectively. Second-site reversions that produced Asn-57.fwdarw. Ile replacements at least partially restored function, presumably by alleviating the instability of these two altered iso-1-cytochromes c. Introduction of the Ile-57 replacement by site-directed mutagenesis in an otherwise normal protein resulted in a 17.degree. C increase in the transition temperature (Tm), corresponding to over a 2-fold increase in the free energy change (.DELTA.G.degree.) for thermal unfolding.