Heteropolymeric Semi-Autonomous Repeat Proteins: Coupling Energetics, Structure, and Function

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Proteins are large molecules that play many critical roles in the body. In this research project, experiments will be performed to learn how protein molecules interact with each other. Proteins often stick together or bind to other proteins of the same type, or of different types, and these types of interactions are critical to protein structure, function, and stability. Though it is fairly easy to "see" these interactions by determining structures of protein complexes, the rules by which the different parts of proteins bind are not known. Here, a series of proteins have been identified in which the protein subunits that bind are all in a single protein chain, making studies of their interactions quite simple to study. Studies will take advantage of the architecture of these "tandem repeat proteins" to learn the rules by which protein subunits stick together. These rules will reveal not only why tandem repeats stick together, but also why separate protein chains stick together. Thus these rules will provide an understanding of protein structure and function in complexes ranging from pairs of protein chains all the way up to the very large protein assemblies that give cells and tissues their shapes and mechanical properties and allow cells to communicate with each other. This research will help train students, both at the undergraduate and graduate level, in the methods of cutting-edge biophysics and biochemistry, both in the laboratory and through the use of computer simulations.These studies will use experimental and computational methods. Fragments of five different proteins will be prepared in the lab, and their structures will be studied with x-ray diffraction and magnetic resonance spectroscopy. Fragment stabilities will be determined using optical techniques like fluorescence and circular dichroism spectroscopy. Computer simulations will also be used to learn what types of interactions promote interaction between protein subunits. A key aspect of this research project is that the repeating structure of proteins studied here can be analyzed with a model called an "Ising" model. This model allows the precise measurement of the strength of interaction between protein subunits. The rules that are learned from this analysis will apply not only to other repeat proteins, but to non-repeating protein assemblies, which use the same types of interactions (packing, charge interactions, and hydrogen bonding) as repeat proteins do.This award reflects NSF's statutory mission and has been deemed worthy of support through evaluation using the Foundation's intellectual merit and broader impacts review criteria.