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13C-based metabolic flux analysis in maize embryos; an approach to identify novel pathways for metabolic engineering

Research Scholar

Mohamed Koubaa, Center for Applied Plant Science (Tunisia)
Ana Paula Alonso, Faculty Mentor

Biography

Mohamed Koubaa earned his engineering diploma in the field of biological engineering at the National Engineering School of Sfax in Tunisia in 2007. He then moved to Compiègne, France where he obtained a master's degree in biotechnology in 2008 and a PhD in 2012. His PhD project was focused on metabolic flux analysis in rapeseed and linseed embryos. He learned of his current supervisor — Ana Paula Alonso, Department of Molecular Genetics — from her publications in the field of plant metabolic flux analysis. He applied for a postdoctoral research position in her lab before his defense, and began his postdoc at Ohio State in April 2012.

What is the issue or problem addressed in your research?

This project specifically addresses major knowledge gaps in biological mechanisms controlling biomass accumulation in seeds, using the highly tractable model system maize (Zea mays). Several maize lines, which differ in biomass content (especially in oil, protein and starch), are commercially available. The aim of this study is to unravel the mechanisms involved in biomass accumulation in maize embryos and gain insight into the identification of possible targets for genetic modifications. The study consists of the development and comparison of carbon flux maps (e.g. between low oil and high oil storage lines) using a powerful tool for metabolic engineering: metabolic flux analysis (MFA). MFA has evolved during the past 20 years as a pivotal concept to describe the metabolic activity of living cells/organs, and to provide the link between observable enzyme activities and metabolite levels.

What methodology did you use in your research?

Maize embryos were incubated with stable isotopic tracers (e.g., [1,2-13C]glucose; [U-13C]glutamine) for 7 days until the isotopic steady state is reached. These tracers were metabolized by cells, resulting in the incorporation of 13C-atoms into intracellular metabolites, macromolecules, and metabolic products. The amount of 13C-labeling and positional distribution of 13C-atoms were then assessed by liquid and gas chromatography – mass spectrometry (LC-MS and GC-MS, respectively), and nuclear magnetic resonance (NMR). Metabolic fluxes were finally calculated from these labeling measurements using a model-based approach that maximizes the fit between the measured and model predicted labeling distributions.

What are the purpose/rationale and implications of your research?

To meet the growing demands of plant bioproducts for food, industrial applications and biofuels, it is imperative to improve crop production. We believe that this project will identify bottlenecks for mechanisms involved in biomass synthesis and accumulation in corn, an important crop in many parts of the world. Redirecting carbon flux towards the production of a compound of interest therefore represents tremendous nutritional and economical benefits. The results from this study will be indispensable to generate new plants that produce higher amount of targeted biomass.