A comparative study of peptide models of the alpha-domain of alpha-lactalbumin, lysozyme, and alpha-lactalbumin/lysozyme chimeras allows the elucidation of critical factors that contribute to the ability to form stable partially folded states

alpha -Lactalbumin (alpha LA) forms a well-populated equilibrium molten glo bule state, while the homologous protein hen lysozyme does not. alpha LA is a two-domain protein and the alpha -domain is more structured in the molte n globule state than is the beta -domain. Peptide models derived from the a lpha -subdomain that contain the A, B, D, and 3(10) helices of alpha LA are capable of forming a molten globule state in the absence of the remainder of the protein. Here we report comparative studies of a peptide model deriv ed from the same region of hen lysozyme and a set of chimeric alpha -lactal bumin-lysozyme constructs. Circular dichroism, dynamic light scattering, se dimentation equilibrium, and fluorescence experiments indicate that the lys ozyme construct does not fold. Chimeric constructs were prepared to probe t he origins of the difference in the ability of the two isolated subdomains to fold. The first consists of the A and B helices of alpha LA cross-linked to the D and C-terminal 3(10) helices of lysozyme. This construct is highl y helical, while a second construct that contains the A and B helices of ly sozyme cross-linked to the D and 3(10) helices of alpha LA does not fold. F urthermore, the disulfide cross-linked homodimer of the alpha LA AB peptide is helical, while the homodimer of the lysozyme AB peptide is unstructured . Thus, the AB helix region of alpha LA appears to have an intrinsic abilit y to form structure as long as some relatively nonspecific interactions can be made with other regions of the protein. Our studies show that the A and B helices plays a key role in the ability of the respective alpha -subdoma ins to fold.