| Amongst Nature's greatest unsolved mysteries is the elusive mechanism by which proteins fold. Current experimental explorations have focused on hydrophobic effects and native like contacts that form in the transition state. In this thesis work, we explore neglected aspects of folding, the contribution of electrostatic and nonnative interaction on the transition state, and the precision of folding measurements. By manipulating ionic strengths of solvent conditions we observe that folding behaviors are well correlated to the theoretical model of Debye-Huckel, facilitating a pure deconvolution of electrostatic contributions. Taking a more focused look at the transition state, non-native interactions were explored through site-specific mutagenesis. We demonstrate that even the most robust negative &phis;-value may not be statistically significant. In this finding we came to ask and explore another question: How precisely can &phis;-values be measured? Performing a multi-lab survey on the same set of proteins we find that &phis;-values are poorly constrained when DeltaDeltaGu is less than 1.7 Kcal/mol. While all the secrets of protein folding have yet to be revealed, much progress has been made on elucidating the details of this complex process. |
