Core B - Biomolecular tools

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CORE B SUMMARY Protein Production and Chemical Biology. The overall goal of this P01 proposal is to elucidate the biological mechanisms of calcification and their potential role in the onset and etiology of age-related macular degeneration (AMD) and Alzheimer’s disease (AD). The proposed projects will investigate calcification at the molecular, cellular and organismal levels to build a comprehensive view of the participants that are responsible for this aging-related phenomenon. Central to all these efforts will be the availability of pure and homogenous protein reagents with defined properties and modifications that can be manipulated in the various assays and experiments proposed in the projects. The goal of this Core will be to design and prepare these reagents, with a particular focus on the calcification-associated proteins vitronectin (Vn), Tau and amyloid β (Aβ). The Core will also aim to develop enhanced tools for the production of modified proteins for structural and imaging studies. More specifically, the Core will work towards the following goals: 1) Production of Vn, Tau, and Aβ constructs for structural, biochemical, biophysical and cellular analysis as necessary for all projects in the proposal; 2) Development of intein-based tools for segmental isotopic labeling of proteins for nuclear magnetic resonance (NMR) studies as described in Project 1; and 3) Preparation of fluorescently labeled proteins for biophysical assays and sensor development for all projects. In addition to serving the projects described in this proposal, we expect that the protocols and reagents developed by the Core will be valuable tools for chemical and cell biologists, biophysicists and structural biologists studying a wide range of biological problems in health and disease.