| The work contained within this thesis focuses on the optical spectroscopic characterization and thermodynamic analysis of selected model beta-sheet peptides, both two-stranded antiparallel beta hairpins and three-stranded antiparallel beta-sheets. Such small peptide systems that have the ability to fold in aqueous solution are invaluable toward understanding beta-sheet folding and stability.; Selected model peptides were utilized to examine the dependence of turn conformation and strand length on beta-sheet formation. Results from FTIR and ECD spectroscopies show that the turn sequence, DPro-Gly, is better for beta-hairpin formation over Asn-Gly. Furthermore, lengthening the beta-strands with Thr, which has a high propensity for beta-sheet structure did not increase beta-sheet content. Thermal denaturation experiments revealed broad unfolding transitions. These transitions could be fit by a two-state model approximation and independently by a statistical mechanical model, which also indicated a two-state system.; The next study involved the estimated quantitation of beta-sheet structure and the thermodynamic characterization of two three-stranded beta-sheet models previously reported by NMR to be highly folded. We found the first model, Betanova, to be mostly unstructured by ECD and FTIR spectra analysis, and the second model, DPDP, to have a substantial amount of beta-sheet structure. Their thermal denaturation profiles displayed nearly linear responses in ECD, FTIR, and fluorescence measurements. Additionally, FRET end labels attached to these peptides confirmed that DP DP is structured in solution providing substantial resonant energy transfer efficiency indicative of the proximity of the N and C termini. Betanova displayed very minor FRET efficiency. FRET efficiency for DP DP as a function of temperature or chemical denaturant concentration was virtually invariant and this was suggestive of a collapsed state for this peptide throughout the unfolding conditions spanned.; Finally, a spectroscopic characterization of the structure and thermodynamic stability of a three-stranded beta-sheet module, the W W domain from hYAP, was conducted. By both ECD and FTIR we determined a beta-sheet content in almost perfect agreement with that obtained from the NMR solution structure. An assessment of the unfolding of this domain revealed that it behaves as a two-state cooperative system for both hydrophobic and interstrand hydrogen bonding interactions. Analysis of truncated constructs of the WW domain indicated that correct folding might require the tertiary context provided by the long (∼10 residues) and unstructured N and C terminal regions. |
