The overall objective of this project is to metabolically engineer the E. coli Sugar-Utilization Regulatory Systems (SURS) to utilize sugar mixtures obtained from plant biomass. The Principal Investigators (PIs) will introduce specific genetic modifications to the SURS and analyze the consequences of these changes at the cellular level. The specific goals are to: (1) construct E. coli strains with modified SURS and evaluate their capability to efficiently utilize sugar mixtures (glucose, xylose and arabinose); (2) evaluate changes in the regulation of gene expression at the genomic scale resulting from engineering E. coli SURS; (3) identify functional metabolic pathways and quantify their fluxes in SURS mutants and wild type strains by using a novel flux analysis technique based on bondomer analysis of 2D NMR bond-labeling experiments; (4) integrate these results using a novel genetic network-based metabolic flux analysis (MFA) model that combines gene expression and metabolic flux data; and (5) propose further genetic/environmental modification for improving the capacity of SURS mutants for fermenting sugar mixtures. The benefits from this project will be two-fold. First, from an applied standpoint, the project will provide a powerful tool for designing of rational modifications of the SURS to obtain recombinant strains capable of efficiently metabolizing sugar mixtures. Second, from a fundamental standpoint, this project will contribute to a better understanding of the role of SURS in regulating both sugar utilization and several other cellular functions.
