Glycosaminoglycans (GAGs), such as heparin, heparan sulfate (HS), and chondroitin sulfate (CS), are naturally occurring polydisperse linear polysaccharides that are heavily O- and N-sulfated. The interaction between GAGs and proteins are critical for many biological processes including cell-cell and cell-matrix interactions, cell migration and proliferation, growth factor sequestration, chemokine and cytokine activation, microbial recognition and tissue morphogenesis during embryonic development. Hundreds of HS-binding proteins have been identified, but the oligosaccharide structure that mediates a particular interaction has been defined in only a few cases due to the structural complexity of HS. In this Technology Research and Development (TR&D2) project, three teams of investigators, representing skills in the areas of nuclear magnetic resonance (NMR), mass spectrometry (MS) and computational biology, will work cooperatively to develop new technologies for the structural characterization of glycosaminoglycan (GAG)-protein complexes and protein-protein complexes that are induced by the presence of GAGs. Their efforts will be guided by the needs of driving biomedical projects (DBPs) that span systems related to cell migration, cell signaling, and the maintenance of cellular integrity. These projects share complexities in the size of the systems involved, the heterogeneity of the interacting GAGs, and the post-translational modification of the proteins. These complexities dictate the development of novel technologies, often in cooperation with other TR&D groups of this Research Resource.
Specific Aims i nclude:
Aim 1. Developing NMR Approaches to Protein-GAG and Protein-Protein Complexes including approaches for identification of bound ligand geometries and interaction motifs and approaches for characterization of protein-protein and GAG-protein complexes, Aim 2. Developing Hydroxyl Radical Protein Footprinting (HRPF) for the structural characterization of Protein-GAG Complexes including technology to improve the sequence coverage of high spatial resolution HRPF, development of HRPF technologies for the analysis of protein-GAG interaction interfaces, and development of HRPF technologies for the analysis of protein-protein interaction interfaces, Aim 3. Developing Computer Modeling approaches for GAGs, Proteoglycans, and GAG-Protein complexes, including optimizing docking algorithms for use with GAGs, adapting the GLYCAM force field and develop suitable protocols to improve energy representation of GAGs
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