BIOPHYSICAL CHARACTERIZATION OF MACROMOLECULES

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The intent of this project is to develop new and improve existing methodology for the characterization of biological macromolecules, and to apply these methods collaboratively to the study of macromolecules and their interactions. Techniques employed are analytical ultracentrifugation, dynamic light scattering, isothermal titration calorimetry, and surface plasmon resonance biosensing. In analytical ultracentrifugation, substantial methodological advances have been made in the modeling of sedimentation velocity data by direct boundary analysis. This includes a novel algebraic noise decomposition method, a least-squares g*(s) method, and the analysis of continuous size distributions. Additional experience was gained in analytical zone centrifugation, a sedimentation velocity method that minimizes the required sample volume. Also, we continued the development of methods for the hydrodynamic characterization of small molecules. We have expanded our expertise in dynamic light scattering and isothermal titration calorimetry, and have written a review on the experimental methods applied in optical biosensing. Experimentally, one major focus of the systems collaboratively studied has been the interactions of membrane receptors and related proteins important in immunology, such as the T-cell receptor, different NK receptors, MHC molecules, superantigen, and FcRn-receptor. A second emphasis was in viral proteins, including HIV-gp120 and herpes simplex capsid proteins. Further systems studied are G-protein subunit interactions, calmodulin interactions with neurogranin, as well as nucleosomes and their interaction with HMG proteins. The characterization of many of these systems has been completed, and several collaborative publications are in press, submitted, and in preparation. - biophysics, binding, thermodynamics, kinetics