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As an approach to learning more about structure-forming processes in chicken egg white lysozyme, the reduced protein was studied in the presence of methanol, 2-chloroethanol, trifluoroethanol, and sodium dodecyl sulfate, at various concentrations in aqueous solution. The mixtures were subjected to gel filtration chromatography, with circular dichroism (CD) examination of the resulting polymers and monomers. Changes in CD behavior, indicating development of ordered structure, were seen to be associated only with polymer formation. Contributing to these changes were increases in beta structure as well as alpha-helix. Since the solutes studied are known as helix formers, it is suggested that studies on many proteins, carried out in the presence of these and other structure-modifying solutes, may have been complicated by formation of beta structure as well as by polymerization. The reduced monomer formed no structure in the hydrophobic environment afforded by these solutes, but only randomized further. Thus, hydrophobic interactions may be of no significance to lysozyme chain folding in its early stages. As an alternative, the long range interactions stabilized by native tertiary cross links may be critical for the transition to native secondary structure.

Gamma radiolysis of dry proteins at 195°K in the presence of tritiated hydrogen sulfide gives information about the distribution of carbon radicals which are precursors of the secondary radicals observed at 298°K. When lyophilized ribonuclease, lysozyme, chymotrypsinogen, and nagarse (a protein without disulfide bridges) are irradiated at 195°K, the observed tritium distributions are approximately equivalent; however, at 298°K characteristic distributions are found. Thus, the mechanisms for the formation of carbon radicals in native proteins can be divided into two groups: first, those which take place below 195°K and are independent of amino acid sequence, conformation, and the presence of disulfide bridges; and second, those which occur above 195°K and are influenced by the specific conformation of each native protein.