Hilty Research Group

Solvent Effect on Polypeptide Structure
Despite an increasing availability of information on the native structure of proteins, our understanding of the molecular mechanisms causing a protein to fold is relatively limited. The difficulty arises due to the presence of a balance between a large number of noncovalent interactions in a polypeptide chain that ultimately determine the protein structure. In addition to intramolecular interactions, a major factor governing protein structure further appears to be solvation effect. Through various solvent-solute interactions, this effect can lead to protein folding or to denaturation.
Obtaining information on the molecular determinants of protein secondary structure formation is greatly facilitated by reducing the complexity of the systems under investigation.
A model system for a soluble protein is the peptide BBA5, which was originally designed to adopt a mixed ββα fold in water. By NMR, the propensity for formation of α-helical and β-sheet secondary structure elements can be addressed separately. Because the identification of specific solvent-solute interactions using NMR spectroscopy alone can be difficult, we collaborate with the Gao Lab at Peking University for performing molecular dynamics (MD) simulations of the same systems.
Additional challenges in identifying interactions leading to protein structure are posed by membrane proteins due to the amphiphilic solvent environment needed for their solubilization. We are studying the folding of disulfide bond forming protein B (DsbB), a membrane protein from the inner membrane of E. coli, which in its native form consists of a bundle of four transmembrane helices. The NMR structures of two peptides corresponding to adjacent transmembrane helices of this protein were determined in different solvents, including helix stabilizing trifluoroethanol, as well as detergent micelle/water mixtures.

Membrane Associated Peptides
Membrane associated α-helices are a category of important structural motifs that are present in all organisms. We are interested in identifying factors that determine the membrane association of small peptides and characterizing the contributions of these factors towards the structure, stability and orientation of these membrane associating helices. Determinants of membrane association of peptides are identified using computational and statistical approaches. Observations of membrane associating peptides in the presence of different membrane mimetics including detergent micelles and organic solvents are carried out using a combination of biophysical methods including circular dichroism and nuclear magnetic resonance spectroscopy and the contributions of these determinants are identified.

Autotransporters
Type V autotransporters constitute a primary protein secretion pathway in Gram negative bacteria. They are very simple in constitution and involve a single protein that can transport itself across the outer membrane. Type V autotransporters have an N-terminal functional domain, which is secreted, and a C-terminal translocator domain that are joined by a linker. The linker region is crucial for the activity of the autotransporter and stability of the barrel. We are using solution NMR techniques to obtain structural and dynamic information that would enable probing the role of the linker in the context of the function of the autotransporter and the folding process of the barrel. We are also investigating the importance of the process of barrel formation and folding in the function of the autotransport process.