Control of the threonine-synthesis pathway in Escherichia coli: a theoretical and experimental approach

Abstract

A computer simulation of the threonine-synthesis pathway in Escherichia coli Tir-8 has been developed based on our previous measurements of the kinetics of the pathway enzymes under near-physiological conditions. The model successfully simulates the main features of the time courses of threonine synthesis previously observed in a cell-free extract without alteration of the experimentally determined parameters, although improved quantitative fits can be obtained with small parameter adjustments. At the concentrations of enzymes, precursors and products present in cells, the model predicts a threonine-synthesis flux close to that required to support cell growth. Furthermore, the first two enzymes operate close to equilibrium, providing an example of a near-equilibrium feedback-inhibited enzyme. The predicted flux control coefficients of the pathway enzymes under physiological conditions show that the control of flux is shared between the first three enzymes : aspartate kinase, aspartate semialdehyde dehydrogenase and homoserine dehydrogenase,

Metabolic Control Analysis of Glycerol Synthesis inSaccharomyces cerevisiae

“…Similarly, in other studies, kinetic models have successfully been used to help researchers understand complex cellular systems. For example, a kinetic model of glycolysis in the bloodstream form of Trypanosoma brucei helped identify factors that affect glycolytic flux (6); similarly, the controlling factors of flux through the threonine synthesis pathway in Escherichia coli have been elucidated (14). Another kinetic model also helped identify targets for manipulation in an effort to maximize the efficiency of conversion of hexose to sucrose and to minimize futile cycling in sugar cane (47).…”

Mathematical model of the Lux luminescence system in the terrestrial bacterium Photorhabdus luminescens

“…The environment of the Lux enzymes in typical experiments is that of an E. coli cell, so the concentrations of ATP, NADPH and FMN found in an E. coli cell were used. 37 The values were 1310 mM for ATP, 560 mM for NADPH, and 88 mM for FMN. The value of the concentration of dissolved oxygen in the cell was not available, so its concentration in water at 37 1C at an atmospheric pressure of 101.3 kPa was calculated,y giving a value of 214 mM.…”

“…37 The analysis identifies the extent to which the control of flux is spread in varying proportions between each of the enzymes and substrates in the system. A flux control coefficient is calculated for each enzyme.…”

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

“…The concentrations for ribulose-5-phosphate (Rb5P) (0.017 mM), R5P (0.062 mM), xylulose-5-phosphate (X5P) (0.022 mM), sedoheptulose-7-phosphate (S7P) (0.025 mM), and E4P (0.020 mM) were adjusted by fitting the product(s) to substrate(s) ratio to be a 0.9 factor of the equilibrium constant (Chassagnole et al, 2002); (5) The concentration of extracellular Glc decayed in the fed batch reactors during the initial 13 h although never below 5 mM (the assumed value), which corresponds to a situation of saturation for the transporter with respect to the substrate; (6) The concentrations of 1,3-diphosphoglycerate (PGP) (0.0025 mM), 2-phosphoglycerate (2PG) (0.11 mM), Asp (4.02 mM), and intracellular Thr (Thr in ) (3.49 mM) were adjusted to fit the flux for PGK, PGMU, ENO, AK, ASD, HDH, homoserine kinase (HK), and TS; and (7) The values of 6-phosphogluconate (6PG) (0.4 mM), aKG (0.50 mM), ACoA (0.20 mM), ASA (0.027 mM), aspartyl-phosphate (AspP) (0.0045 mM), fumarate (Fum) (0.1 mM), HS (0.037 mM), O-phosphohomoserine (HSP) (0.041 mM), Lys in (0.46 mM), NAD þ (1.47 mM), NADH (0.1 mM), NADP þ (0.20 mM), NADPH (0.18 mM), OAA (0.03 mM), and inorganic phosphate (Pi) (5 mM) were selected to match the values found in the literature (Albe et al, 1990;Peng et al, 2004) or these applied in the original models (Chassagnole et al, 2001b(Chassagnole et al, , 2002. Lys in refers to intracellular lysine.…”