Developing Proteomic Approaches for DNA-protein Interactome Discovery and Global Proteoform Elucidation
Author | : Yunxiang Dai |
Publisher | : |
Total Pages | : 0 |
Release | : 2019 |
Genre | : |
ISBN | : |
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Proteins exert essential molecular functions in numerous biological processes. Mass spectrometry-based proteomics has developed into a powerful approach to analyze proteins and study protein functions at large scale in a complex system. From a proteomic perspective, functional investigation of proteins aims to elucidate proteins' comprehensive interaction networks, as well as their structural specificity that fulfills the various biomolecular activities. Development of proteomic platforms to tackle particular protein function questions requires all stages to work cohesively. These include careful isolation of the proteome of interest from complicated biological samples, effective analysis within liquid chromatography/mass spectrometry instrumentation, and thorough examination of high-throughput proteomic data. In the works presented, I advanced proteomic approaches to probe two intriguing topics associated with protein function: DNA-protein interaction and global intact proteoforms. In the strategy targeting the former subject, shown in Chapter 2, the HyCCAPP (Hybridization Capture of Chromatin Associated Proteins for Proteomics) method was optimized to allow extraction of multiple genomic loci and their interacting proteins in parallel, followed by identification using bottom-up mass spectrometry. Chapters 3 and 4 elaborate strategies to identify global proteoforms with specific post-translational modification information in both prokaryotic and mammalian systems. The innovative analytical approaches integrated different schemes including intact-mass, bottom-up, and top-down proteomics for enhanced identification and construction of proteoform families. The technologies demonstrated in this dissertation will hopefully continue to evolve to accommodate a multitude of studies in functional proteomics, and provide insights into unsolved problems in health and disease-related systems biology.