In silico strategy to rationally engineer metabolite production: A case study for threonine in Escherichia coli

Abstract

Genetic engineering of metabolic pathways is a standard strategy to increase the production of metabolites of economic interest. However, such flux increases could very likely lead to undesirable changes in metabolite concentrations, producing deleterious perturbations on other cellular processes. These negative effects could be avoided by implementing a balanced increase of enzyme concentrations according to the Universal Method [Kacser and Acerenza (1993) Eur J Biochem 216:361-367]. Exact application of the method usually requires modification of many reactions, which is difficult to achieve in practice. Here, improvement of threonine production via pyruvate kinase deletion in Escherichia coli is used as a case study to demonstrate a partial application of the Universal Method, which includes performing sensitivity analysis. Our analysis predicts that manipulating a few reactions is sufficient to obtain an important increase in threonine production without major perturbations of metabolite concentrations.

unIn silico Derivation of a Reduced Kinetic Model for Stationary or Oscillating Glycolysis in Escherichia coli Bacterium

“…Various applications are reported, such as optimization of synthesis of amino-acids (AA), [19][20][21][22] ethanol, [23][24] succinate, 25 lactate, 26 or other metabolites in mutant cells 27,28 . Deterministic simulation platforms allow in silico design of modified cells with desirable gene circuits and 'motifs' of practical applications in the biosynthesis industry, environmental engineering, and medicine.…”

A modular modulation method for achieving increases in metabolite production

“…The aim is, therefore, to develop approximate solutions, achieving similar results and requiring a smaller amount of work . It has been shown with a model of an E. coli overproducing strain that manipulating a small number of the steps involved in the universal method may be sufficient to obtain a solution relatively close to the exact one. The MMM, developed in this work, was devised to determine, experimentally, where those key steps for manipulation are located in the network.…”

“…In many cases, the exact application of the universal method requires changing a relatively high number of steps, which may be difficult to implement in practice. However, model simulation has shown that, of all the steps corresponding to the exact application of the method, it may be sufficient to manipulate only a small number of key steps to obtain the desirable flux increase . Strategies, previously developed, to determine these key steps are limited by restrictive assumptions.…”

Effects of Cadmium and Mercury on the Upper Part of Skeletal Muscle Glycolysis in Mice

“…The simultaneous decrease of the limiting rates of HK and PFK, the enzymes controlling the flux, is equivalent to a minimal implementation with only two enzymes of multisite modulation, which is an efficient way by which organisms achieve very large flux changes, but with only small concentration changes in internal metabolites [37], [38], [39]. Fig.…”