Abstracts Chemistry

Characterizing the unfolding pathway and denatured state of proteins through molecular dynamics simulations

by Steven Lee Kazmirski

The denatured state of proteins is typically assumed to contain no structure. However, recent years have produced experimental studies that argue that there is residual structure in the denatured state of proteins. This residual structure can be important to the folding and stability of the protein. Obtaining an atomic resolution view of the denatured state is currently impossible through experimental means, and molecular dynamics simulations have been used to unfold proteins and observe the unfolding pathway.In this thesis, the first three chapters each discuss multiple unfolding simulations of different proteins. Chapter 1 discusses the unfolding of bovine pancreatic trypsin inhibitor (BPTI) and the residual structure in the denatured state. The ensemble of denatured structures have been compared back to specific distances defined by fluorescence and NMR experiments. Chapter 2 describes two 9 ns unfolding simulations of hen eggwhite (HEW) lysozyme. Previous simulations of the unfolding pathway of HEW lysozyme from other laboratories never agreed correctly with the experiments monitoring the unfolding of the two domain of lysozyme. The simulations in this thesis do unfold correctly with the beta-domain unfolding before the alpha-domain. Also, five intermediates were defined and their non-native structure is defined. Chapter 3 defines the residual structure in chymotrypsin inhibitor 2 (CI2). The unfolding of CI2 is two-state experimentally. Previous simulations and experiments have characterized the native and transition states. The simulations in chapter 3 fill in the last part of the folding pathway, showing the little residual structure in the denatured state.The final three chapters move away from the denatured state of specific proteins. Chapter 4 is a discussion on new analysis techniques for comparing multiple unfolding trajectories. These new techniques borrow on techniques used in other fields such as principal component analysis. Chapters 5 and 6 describe the effect of mutations on a peptide from the prion protein. This peptide is believed to be important in the transition of the normal cellular form of the protein to its infectious scrapie form.