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Genome-scale metabolic models are an invaluable tool for analyzing metabolic systems as they provide a more complete picture of the processes of metab. The authors have constructed a genome-scale metabolic model of Escherichia coli based on the iJR904 model developed by the Palsson Lab. at the University of California at San Diego. Group contribution methods were utilized to est. the std. Gibbs free energy change of every reaction in the constructed model. Reactions in the model were classified based on the activity of the reactions during optimal growth on glucose in aerobic media. The most thermodynamically unfavorable reactions involved in the prodn. of biomass in E. coli were identified as ATP phosphoribosyltransferase, ATP synthase, methylene-tetrahydrofolate dehydrogenase, and tryptophanase. The effect of a knockout of these reactions on the prodn. of biomass and the prodn. of individual biomass precursors was analyzed. Changes in the distribution of fluxes in the cell after knockout of these unfavorable reactions were also studied. The methodologies and results discussed can be used to facilitate the refinement of the feasible ranges for cellular parameters such as species concns. and reaction rate consts. [on SciFinder (R)]