Engineering a living cell to desired metabolite concentrations and fluxes: pathways with multifunctional enzymes

With molecular genetics enabling modulation of the concentrations of cellular enzymes, metabolic engineering becomes limited by the question of which modulations of the enzyme concentrations are required to bring about a desired pattern of cellular metabolism. In an earlier paper (Kholodenko et al. (1998). Biotechnol. Bioeng. 59, 239-247) we derived a method to determine the required modulations. This method, however, cannot be immediately applied to cellular pathways with enzymes catalyzing more than one step in metabolism (multifunctional enzymes). In the present paper we show to which extent the presence of multifunctional enzymes limits biotechological ambitions, which one might otherwise pursue in vain. In particular, it is impossible to change the concentration of a single intermediate and leave the rest of metabolism unperturbed if that intermediate interacts directly with a multifunctional enzyme. The analytical machinery of Metabolic Control Analysis is used to relate the desired and ensuing changes in the metabolic pattern. An explicit solution to this problem of engineering metabolism is then given in the form of a single matrix equation.