Elucidation of Pathways: Bacterial and Mass Spectrometric Studies

Bacteria are intimately associated with the human body; beneficial bacteria support health and well-being by aiding digestion, protecting against infection, and stimulating the immune system. Substances produced by bacteria, known as metabolites, act as molecular signals defining the bacterium-host relationship. To potentially benefit the quality of life by revealing new directions for the treatment of disease, our approach is to thoroughly study Fusobacterium varium, a bacterium typically found in the intestines of animals that may contribute to the initiation of bowel disease. Our research program also focuses on chemical processes associated with mass spectrometry, a technique in which molecules are first charged and then broken into pieces upon input of energy. The specific pieces formed are diagnostic, providing a basis for the identification of drugs, environmental contaminants and explosives in various situations and allowing conclusions to be drawn that have broad implications (e.g., the use of banned substances in sports). To develop a deeper understanding of fundamentals behind these important applications, we choose substances that mimic the structures of current drugs and drug candidates, and can accommodate charge in more than one location, increasing their likelihood to break apart in two or more different ways (i.e., fragmentation pathways). With our interpretative developments, mass spectrometry will provide enriched and more reliable information, increasing its value as an essential tool for the characterization of drug metabolites and shortening the time needed to complete this essential step in the development of prospective new drugs. As a complement to genetic studies, our on-going chemical research has demonstrated metabolic diversity in the intestinal bacterium leading to the production of eight substances, of which one may have an unanticipated function as a molecular signal with implications in bowel disease. By identifying the pathways used to form these metabolites, unique metabolic processes in anaerobic bacteria can be evaluated as targets for selective metabolic inhibitors, a novel rational design strategy for the generation of lead compounds for novel antibiotics effective against pathogenic bacteria resistant to current antibiotic treatments. Also, three of the bacterial metabolites excreted have important practical applications as sustainable chemical feed stocks used, for example, in the manufacture of biodegradable polymers with biomedical applications. In this application, funding is requested to pursue two seemingly diverse research investigations. However, commonalities in the thought process of our mechanistic investigative approach to pathway elucidation and the nature of the substances studied provide significant overlap. For each, we design and synthesize probes to test pathway hypotheses, collect evidence using substances labelled with stable isotopes, and logically assimilate the evidence to provide a reliable description of each pathway. Overall, we tackle fundamental questions in the relationship between intestinal bacteria and human hosts, a complex and relatively unexplored area at the intersection of bacterium and human metabolism. Our research program in fundamental science will offer new insights for the interpretation and application of important sequential consecutive chemical processes, the implementation of sustainable fermentation methods, and potentially new avenues of drug research and discovery.