Towards kinetic modeling of genome-scale metabolic networks without sacrificing stoichiometric, thermodynamic and physiological constraints

Mathematical modeling is an essential tool for the comprehensive understanding of cell metabolism and its interactions with the environmental and process conditions. Recent developments in the construction and analysis of stoichiometric models made it possible to define limits on steady‐state metabolic behavior using flux balance analysis. However, detailed information on enzyme kinetics and enzyme regulation is needed to formulate kinetic models that can accurately capture the dynamic metabolic responses. The use of mechanistic enzyme kinetics is a difficult task due to uncertainty in the kinetic properties of enzymes. Therefore, the majority of recent works considered only mass action kinetics for reactions in metabolic networks. Herein, we applied the optimization and risk analysis of complex living entities (ORACLE) framework and constructed a large‐scale mechanistic kinetic model of optimally grown Escherichia coli. We investigated the complex interplay between stoichiometry, thermodynamics, and kinetics in determining the flexibility and capabilities of metabolism. Our results indicate that enzyme saturation is a necessary consideration in modeling metabolic networks and it extends the feasible ranges of metabolic fluxes and metabolite concentrations. Our results further suggest that enzymes in metabolic networks have evolved to function at different saturation states to ensure greater flexibility and robustness of cellular metabolism. Using mass action kinetics in modeling metabolic processes can lead to overly conservative assessments of feasible physiological conditions of the organism. This study shows that considering the enzyme saturations as observed in biological systems consistently increases the stability of the kinetic models. This indicates that enzymes have evolved in a way to increase the flexibility and thus the viability and adaptability of the living organism. Thus, for consistent modeling of cellular metabolism enzyme saturation should be considered as an inevitable component.