Characterization of a peptide model for protein unfolded states. Trevor P. Creamer, Department of Molecular and Cellular Biochemistry, University of Kentucky The process by which proteins adopt their unique functional shapes, or structures, is known as protein folding. Understanding this process has long been one of the more interesting, important, and vexing problems in biophysical chemistry. It has become clear that the single biggest barrier to an understanding of the folding process is a lack of information on where it starts - protein unfolded states. We have been studying a unfolded state model system that consists of short peptides (fragments of proteins). The underlying base or host peptide has been designed to have understandable structural tendencies, but is too small to fold into a stable structure. This host peptide, called A, adopts an ensemble of structures with clear -helix content. We have substituted a variety of guest residues into this host peptide A and studied how they perturb the ensemble of conformations. Using circular dichroism spectroscopy, we have found the unfolded peptide ensemble is dominated by two conformations, the -helix and the polyproline II (PII) helix. The equilibrium between these states changes as a function of the identity of the guest residue and in response to solution conditions. -Helix PII Helix Other structures Our major finding so far is that the well-established propensities for residues to be part of -helices are poor predictors of unfolded state behavior. Our continued studies of this, and other models, for protein unfolded states will begin to give us a better sense of where protein folding starts. This will enable us to better understand the folding process and bring us closer to solving this important problem. CD spectra demonstrating response of peptide A to TFE, an -helix inducing co-solvent, and urea, which induces PII helix. CD spectra of host peptide, A, plus peptides with -helix tolerant guests Q and L, and PII-inducing guests G and P.