Development of metabolic flux and control analysis for enhanced lycopene production in escherichia coli

Abstract

Recently metabolic engineering has concerned systematic strategies to investigate the rational design for strain improvement through genetic manipulations of cellular metabolism by recombinant DNA technology. Production of lycopene (C40), used as an anti-oxidant and anti-cancer agent, in microorganisms has been focused to replace the lycopene production from the extraction of reddish plants. However, the systematic approaches for the improvement of the yield in microorganisms were not developed yet. In this study, I developed two strategies for enhanced lycopene production in Escherichia coli through the metabolic flux and control analysis. First, flux scanning based on enforced objective flux (FSEOF) was the algorithm of the selection of the several candidates for the gene amplification on genome-scale metabolic model to improve the yield of the desired product. FSEOF was applied to enhance lycopene production in E. coli. It led that a twofold (6.48 mg/gDCW) over parental strain and a 6-fold over wild type strain of E. coli. Second, dynamic metabolic control analysis was applied to investigate the shifted rate-controlling steps after the role of the first rate-controlling step was reduced. As a result, phosphofructokinase and pyruvate kinase could be considered as the positive and negative potential rate-controlling enzymes, respectively in lycopene production in E. coli. The amplification of pfk gene from L. bulgaricus in pykFA-knockout mutant had over 50% improvement of lycopene content in E. coli. Based on the metabolic flux and control analysis, the developed systematic approaches for strain improvement can be extended to other valuable product in various microorganisms.