An extended and generalized framework for the calculation of metabolic intervention strategies based on minimal cut sets

Abstract

The concept of minimal cut sets (MCS) provides a flexible framework for analyzing properties of metabolic networks and for computing metabolic intervention strategies. In particular, it has been used to support the targeted design of microbial strains for bio-based production processes. Herein we present a number of major extensions that generalize the existing MCS approach and broaden its scope for applications in metabolic engineering. We first introduce a modified approach to integrate gene-protein-reaction associations (GPR) in the metabolic network structure for the computation of gene-based intervention strategies. In particular, we present a set of novel compression rules for GPR associations, which effectively speedup the computation of gene-based MCS by a factor of up to one order of magnitude. These rules are not specific for MCS and as well applicable to other computational strain design methods. Second, we enhance the MCS framework by allowing the definition of multiple target (undesired) and multiple protected (desired) regions. This enables precise tailoring of the metabolic solution space of the designed strain with unlimited flexibility. Together with further generalizations such as individual cost factors for each intervention, direct combinations of reaction/gene deletions and additions as well as the possibility to search for substrate co-feeding strategies, the scope of the MCS framework could be broadly extended. We demonstrate the applicability and performance benefits of the described developments by computing (gene-based) Escherichia coli strain designs for the bio-based production of 2,3-butanediol, a chemical, that has recently received much attention in the field of metabolic engineering. With our extended framework, we could identify promising strain designs that were formerly unpredictable, including those based on substrate co-feeding.

Increasing ATP turnover boosts productivity of 2,3-butanediol synthesis in Escherichia coli

“…For applications of enforced ATP wasting, obligate coupling, where balanced ATP synthesis is only feasible with product formation, is the ideal case. Computational strain design methods have been developed to identify intervention strategies that lead to such obligate coupling of ATP (or, alternatively, biomass) synthesis with product formation [ 36 , 37 ]. One possible way to achieve strong coupling of ATP and 2,3-BDO synthesis is to use microaerobic instead of fully aerobic conditions where, on the one hand, the NADH supply for 2,3-BDO synthesis increases due to limited oxygen availability, while the amount of supplied oxygen exactly balances the remaining surplus of NADH when producing 2,3-BDO from glucose.…”

Designing microbial communities to maximize the thermodynamic driving force for the production of chemicals

“…Finding suitable knockout strategies that leave only the relevant pathway segments active in the respective strains can be computed by dedicated strain design algorithms, e.g. based on minimal cut sets [ 51 ]. Implementing the respective gene knockout strategies may lead to unstable strains or even prevent growth.…”

Environmental flexibility does not explain metabolic robustness

“…However, we have demonstrated previously that PoF analyses at the level of genes and at the level of reactions yield qualitatively similar results 13 . Incidentally, recent advances in software for the enumeration of genetic MCSs 62 , 63 allow for extending the scope of our method to the gene level. Additionally, we have shown here that the PoF is independent of the network’s size and mostly determined by the number of essential reactions (or genes, for that matter).…”