Development and improvement of systemic approaches for the enhanced production of metabolites based on the metabolic flux analysis

Abstract

The several systematic approaches in metabolic engineering have been successfully applied to the improvement of microbial strains for the overproduction of desired bioproducts. However, various in silico methodologies for systemic analysis are necessary to be improved, which allow more precise data set and another prediction strategy for strain improvement, which is successful strategy identifying genes for the flux amplification. No doubt that the development of systematical strategy for the identification of genes responsible for the improvement of producing strain by the genetic amplification will be challengeable in functional genomics era. In here, we report a new strategy of combining the advantages of constraints-based metabolic flux analysis and $^{13}C$ -constrained flux balancing analysis. The new strategy allows more accurate determination of metabolic fluxes by employing the mass balances around artificial metabolites and the converging ratio determinants (CRDs) as additional constraints. Furthermore, this improved methodology allows the pathway analysis of metabolic network model with some experimental constraints. In addition, we developed flux scanning based on enforced objective flux (FSEOF) based on constraints-based analysis with genome-scale model to select gene targets for the amplification of fluxes. In application, we presented the examples of strain improvement for enhanced production of lycopene, poly(3-hydroxybutyrate), shikimic acid, and recombinant protein in E. coli by using the gene amplification for the predicted genetic targets identified by FSEOF.