Manley Hoppe Professor

My current research interests are in the areas of plant and microbial metabolic engineering, particularly in the analytical tools for measuring fluxes and concentrations in metabolic pathways. All projects in my laboratory are in collaboration with plant scientists and biological engineers in a portfolio of metabolic engineering applications: overproduction of valuable plant natural products in hairy roots, synthesis of hydrocarbons in plants and algae, protein and oil production in soybean embryos, and production of biorenewable chemicals from E. coli and S. cerevisiae. My laboratory focuses on the metabolic characterization of the biological system in the ME application and uses several techniques to analyze metabolites, nutrients and fluxes. These include ¹³C NMR and MS techniques to monitor primary carbon metabolism, HPLC and LC/MSn to measure secondary metabolites.

• Metabolic Engineering for Biorenewable Chemicals: Our team of interdisciplinary researchers from CBiRC is designing microbes that can overproduce biorenewable chemicals. A key challenge for the commercial production of biorenewable chemicals is to shorten the metabolic engineering design cycle time for the development of high yielding microbial biocatalysts. Metabolic flux analysis is a key component of this design process. An example of an ongoing collaborative project involves the creation of an integrated flux platform technology. An integrated flux platform technology uses comprehensive experimental flux analysis to mathematically constrain an in silico metabolic model of the microbe, and then computationally predicts the complete set of genetic modifications leading to the overproduction of the target chemical. The genetic interventions are prioritized computationally based on their impact on product yield and ordered in a logic chain. Our ultimate goal is to develop robust integrated flux platform tools that will, in turn, accelerate the commercialization of microbial-based technologies for the efficient production of biorenewable chemicals.
• Phytochemical Engineering: The powerful anticancer agents vinblastine and vincristine are obtained commercially from the intact plants of the periwinkle Catharanthus roseus. The low yield of these valuable indole alkaloids in plants has been the major motivation to produce them by cell and tissue cultures. We are interested in the metabolic engineering of the terpenoid indole alkaloid pathways for the overproduction of valuable alkaloids transgenic C. roseus “hairy root” cultures. Flux of carbon into the alkaloid pathways, diversion of flux at intermediate branches, and lack of final conversion at the end of a specific branch all appear to affect alkaloid production. Precise genetic modification of the pathways and subsequent metabolic analysis of fluxes are enabling the identification of bottlenecks in the “working model” of the pathways. By identifying points of flux limitation, pathway steps then can be pursued for genetic modification in a reiterative process, or if the genes have not been cloned, further studies can be targeted to obtain the unknown information.
• Metabolic Flux Maps in Plants: Metabolic flux analysis quantifies the rate of carbon flow for each metabolic reaction in a biochemical pathway model. The approach requires formulation of balanced equations around each metabolite in the network. These metabolite balances are complemented with extracellular measurements of substrate consumption, secretion of metabolites, biomass composition and intracellular measurements such as ¹³C labeling data detected using nuclear magnetic resonance (NMR) spectroscopy or mass spectroscopy (MS). Application of ¹³C labeling-based metabolic flux analysis towards understanding plant physiology has been limited due to the mathematical burden associated with solving a complex model that accounts for comprehensive and rigorous analysis of the NMR data in addition to cellular compartmentation. We have developed a comprehensive generic mathematical tool (NMR2Flux) for metabolic flux analysis that provides network topology information and quantitatively determine fluxes in different cellular compartments. We have applied the metabolic flux map tool in a variety of systems, including soybean embryos, maize cell suspensions, and C. roseus hairy roots.