REFOLDING OF RNASE-A AT HIGH-CONCENTRATIONS - IDENTIFICATION OF NONNATIVE SPECIES

In this paper, we present an analysis of the soluble species formed on refolding of RNase A at various concentrations, in order to character ize these species with respect to structure and activities. Studies we re carried out using reverse-phase high-performance liquid chromatogra phy, circular dichroism, chromatography and ultracentrifugation. At al l concentrations of protein used, RNase A refolded to the native form, together with formation of non-native species. These non-native speci es are either misfolded monomers or aggregates; the percentage of such species increases with increasing concentration of enzyme. Such aggre gation appears to be a non-random process governed by intermolecular d isulfide crosslinking between monomers. These results reaffirm the pri nciple that the information for folding of the protein is encoded in t he amino acid sequence itself.