Metabolic Engineering in Biomedicine

Plant Metabolic Engineering Under Construction: Design Rules and Tools Needed

Jacqueline V. Shanks

Iowa State University

Objective:

To engineer C. roseus hairy roots for the overproduction of tryptophan and indole alkaloids

Approach:

	Develop an inducible promoter system in Catharanthus roseus hairy root cultures
	Construct and characterize transgenic hairy root lines overexpressing key enzymes in the indole and non-mevalonate terpenoid pathways
	Develop methods for NMR-based metabolic flux maps
	Perform metabolic characterization of these first generation transgenic lines
	Develop techniques for co-transformation that allow the introduction of multiple genes
	Construct and characterize a second generation of transgenic lines expressing strategic combinations of genes as determined from the results of the first generation transgenic lines

Accomplishments:

	Glucocorticoid-mediated inducible promoter system developed in Catharanthus roseus hairy root cultures
	Development of the bondomer concept and software in analysis of NMR carbon-bond labeling experiments
	Demonstration of an NMR flux map for central and intermediary carbon metabolism in a three compartment model (cytoplasm, mitochondrion, and plastid) in plant tissues
	NMR flux map tool extended to crop plants (soybean)
	Huge overproduction of tryptophan (>300-fold) and tryptamine (>10-fold) in engineered hairy root lines and small increases in indole alkaloid production

Impact:

	Enhance the nutritional value of crops for human and animal consumption
	Enhance the overproduction of medicinal metabolites in crops

Abstract:

Plants are the raw materials for the world’s feed supply as well as biobased industrial products. Providing an ample food supply for all humankind is a serious challenge for our society. Plants as a biorenewable resource have the potential for providing society with many of our basic goods as well as energy. The design of plants to meet these potentially competing demands is a scientific and engineering challenge. Plant metabolic engineering provides a means of understanding plant biology - “systems biology”. Plant metabolic engineering is also the basis of designing new plants. “Predictive metabolic engineering” (Sweetlove, Last, and Fernie, 2003) is a worthy goal for both aspects. The integration of knowledge for prediction will require design software as well as powerful experimental tools. While comprehensive integration of “omics” data in a mathematical model is an ideal goal, using these types of techniques on subsets of metabolism could help develop logic-based design rules, instead of empirical ones. Plants, by both their complexity and timescale of growth, impose formidable requirements for quantitative analysis and precise synthesis tools. This talk will highlight some of our work in engineering of overproduction of tryptophan and indole alkaloids in hairy root cultures of the medicinal plant Catharanthus roseus.

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