Browsing Theses and Dissertations School of Pharmacy by Subject "Receptors, Pattern Recognition"
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Advances in Mass Spectrometric Structural Biology Techniques for Pattern Recognition Receptor Ligands of Microbial OriginPattern recognition receptors (PRR) are the innate immune system’s first-line sentinels for distinguishing “self” from “non-self.” Many molecules found in the bacterial cell wall are PRR ligands, including lipopolysaccharide (LPS), cardiolipin (CL), and peptidoglycan (PGN). Molecular structural biology techniques are essential for determining both basic cell biology and host-bacteria interactions through ligand-receptor binding mechanisms. Recent interest in designer PRR ligands or PRR ligand mimetics for use in drug discovery pipelines have given this research more translational value as well. Mass spectrometry (MS) has the unique capability to derive primary structures of ions as well as monitor many different ions in complex mixtures. Several different advances in PRR ligand structure analysis were achieved in this dissertation. First, chemical structure of an LPS-derived vaccine was determined using a top down tandem MS approach. Several different instrumental configurations and methods were employed to demonstrate complementarity of data and broad applicability of the approach. Second, CL from a newly discovered Actinomycete marine sponge symbiont was analyzed and compared to CL from a terrestrial Firmicute to generate hypotheses about host-bacterium interactions. This was the first molecular analysis of any secondary metabolites from this species of bacteria. Third, PGN subunits (muropeptides) from Rickettsia typhi were analyzed in a data dependent global LC-tandem MS approach. This was the first example of PGN structure discovery for R. typhi and the first example of this approach applied to PGN structure elucidation for any Rickettsiae species. All of these developments will help to advance PRR-ligand interaction research – an emerging and promising field for development of novel disease treatment and prevention approaches. Modulation of the innate immune response to bacterial insult is a challenging task without a clear understanding of underlying molecular mechanisms and how they might be manipulated by medicine. One key step in this process is development of sensitive and specific chemical analysis methods fit to acquire unequivocally interpretable data. While all of the methods described herein were applied to specific biological problems, their applicability to other scientific questions is broad.