RII Track-4: Using in-cell NMR to follow 13C-fluxomics in living cells

Boise State University, Boise ID

Investigators

Abstract

Non-technical Description Metabolic pathways in living systems are complex networks of chemical reactions that convert small molecules, known as metabolites, into fundamental energy and chemical building blocks, such as proteins, lipids, and nucleic acids. The study of metabolites produced in living cells and tissues is broadly termed 'metabolomics'. Metabolomics can provide information on how changes in a cell?s environment impacts those complex biochemical networks. Monitoring the concentrations of metabolites over time can provide essential information about the health of an organism. Metabolomics is emerging as an integral technology to understand the function of a wide variety of biological systems in response to a range of conditions, including disease, drug treatment, and exposure to pests or pathogens. The PI will work with colleagues at the National Renewable Energy Lab (NREL) to answer fundamental questions about the flow of metabolites in bacterial cells, which will broadly impact areas of research where understanding basic cellular metabolic processes are important. The project will not only improve the PI's research trajectory, but is also likely to result in significant advances in methods used to study metabolic processes in industrially and environmentally important bacteria. This project will provide interdisciplinary training and research opportunities for undergraduate and graduate students at Boise State University and regional high school students from demographically diverse backgrounds. Technical Description The goal of this project is to decipher carbon utilization in the metabolic pathways of two strains of live fermenting bacteria, Clostridium ljungdahlii and Clostridium thermocellum, both available at NREL. Achieving this goal will provide insights into pathway reversibility and carbon flux at the systems level. Information gained from this study will guide rational bioengineering efforts to optimize biofuel and chemical product output, and increase carbon utilization efficiency. The overarching strategy of this project is to combine advanced in-cell nuclear magnetic resonance (NMR) with 13C-isotope enriched metabolic tracers and paramagnetic relaxation enhancement (PRE) to measure intra- and extracellular metabolic flux (13C-fluxomics) in these microbes. The proposed research will: (1) expand basic science knowledge of metabolic flux in living systems across the molecular, subcellular, and cellular scales using in-cell NMR; and (2) unlock deep fundamental insights into the cellular bioenergetics of fermenting biomass while mitigating CO2 production. This research integrates several novel approaches that will allow real time analysis of 13C-fluxomics.