This project entails structure and dynamics studies of biological macromolecules through the use of nuclear magnetic resonance (NMR) spectroscopy. The present foci are polysaccharides, important vaccine components whose structure is poorly understood, and the structure of Interferon (IFN) -alpha 2c and alpha-21a and their hybrids. I am using NMR to detect changes in the spectra of carbohydrates such as sucrose, cellobiose, maltose and lactose. These simple disaccharides will serve as models to provide the basis for structural studies of oligo- and polysaccharides. To date, I have found that dissolving carbohydrates in liquid crystalline media orients them in a magnetic field, and this orientation provides important and direct structural data, which previously had to be inferred. Specifically, I am measuring inter-nuclear residual dipolar interactions, and these interactions allow me to relate atoms to one another. One interesting outcome of the preliminary work is the suggestion that the two rings in sucrose are perpendicular. The solution structure of sucrose is still hotly debated, and these results will help answer important questions in this debate. Established methodology for measuring residual dipolar couplings yielded inaccurate results for carbohydrates. Consequently we are developing new methodology which will provide accurate measurements of inter-nuclear dipolar interactions. Upon proving this methodology robust in model systems, we will study polysaccharide structure in a similar fashion. I have also verified, using C-13 nuclear relaxation measurements, that the liquid crystals and the disaccharides don't interact significantly, hence the liquid crytals are not interfering with the measurements of residual dipolar couplings. Another project is the solution structure of IFN hybrids. Our goal in this work is to characterize the structure of the parent proteins (IFN-alpha 2c and IFN-alpha 21a), and their hybrids and use them to understand the interactions which affect their anti viral and anti proliferative acitivities. IFN alpha is approved for the treatment of AIDS-related Kaposi's Sarcoma and for Hepatitis B and C. The goal of this project is to characterize the structure of IFN hybrids by multidimensional solution NMR spectroscopy and to use these structures to understand structure-function relationships. A deeper understanding of IFN structure could lead to the design of mroe effective therapeutics with lower toxicity.
