Elucidation and structural analysis of conserved pools for genome-scale metabolic reconstructions

Abstract

In this article, we introduce metabolite concentration coupling analysis (MCCA) to study conservation relationships for metabolite concentrations in genome-scale metabolic networks. The analysis allows the global identification of subsets of metabolites whose concentrations are always coupled within common conserved pools. Also, the minimal conserved pool identification (MCPI) procedure is developed for elucidating conserved pools for targeted metabolites without computing the entire basis conservation relationships. The approaches are demonstrated on genome-scale metabolic reconstructions of Helicobacter pylori, Escherichia coli, and Saccharomyces cerevisiae. Despite significant differences in the size and complexity of the examined organism's models, we find that the concentrations of nearly all metabolites are coupled within a relatively small number of subsets. These correspond to the overall exchange of carbon molecules into and out of the networks, interconversion of energy and redox cofactors, and the transfer of nitrogen, sulfur, phosphate, coenzyme A, and acyl carrier protein moieties among metabolites. The presence of large conserved pools can be viewed as global biophysical barriers protecting cellular systems from stresses, maintaining coordinated interconversions between key metabolites, and providing an additional mode of global metabolic regulation. The developed approaches thus provide novel and versatile tools for elucidating coupling relationships between metabolite concentrations with implications in biotechnological and medical applications.

FIGURE 1

A metabolite subset that comprises metabolites A, B, and C simultaneously present in conserved pools 1, 2, and 3.

FIGURE 2

Example representation of a cellular system in the context of complete genome-scale metabolic reconstructions. A complete metabolic reconstruction encompasses both internal (i.e., B, C, E, and F) and external (i.e., A and D) metabolites and thus the corresponding stoichiometric matrix describes a closed system. Here V₁ and V₃ are transport reaction fluxes across the cellular system's boundary, V₂ and V₄ are intracellular reaction fluxes, and W_A and W_D are external exchange fluxes.

FIGURE 3

Extreme pools identified for glycolysis. Reactions are: 1), hexokinase, 2), phosphoglucoseisomerase, 3), phosphofructokinase, 4), fructose 1,6-bisphosphatase, 5), aldolase, 6), triose phosphate isomerase, 7), glyceraldehyde 3-phosphate dehydrogenase, 8), phosphoglycerate kinase, 9), phosphoglycerate mutase, 10), enolase, 11), pyruvate kinase, 12), ATPase, and 13), adenylate kinase. Metabolites are: glucose (Glu), glucose 6-phosphate (G6P), fructose 6-phosphate (F6P), fructose 1,6-bisphosphate (F1,6P), dihydroxyacetone phosphate (DHAP), glyceraldehyde 3-phosphate (G3P), 1,3-bisphosphoglycerate (1,3BPG), 3-phosphoglycerate (3PG), 2-phosphoglycerate (2PG), phosphoenolpyruvate (PEP), pyruvate (Pyr), and inorganic phosphate (P_i).

FIGURE 4

The various potential types of coupling between two metabolites are related to their conservation coefficient ratio limits R_min and R_max as shown. Two metabolites are: 1), fully coupled if the presence of one metabolite within conserved pools implies the presence of the other within the same conserved pools and vice versa, 2), partially coupled if they are always present within common conserved pools but their conservation coefficients are not fixed, and 3), directionally coupled if the presence of one metabolite within conserved pools implies the other and not vice versa.

FIGURE 5

A schematic representation of couplings between metabolites. Metabolites A and B are fully or partially coupled, and are simultaneously present in all common conserved pools (i.e., pools 1, 2, and 3). These metabolites form a metabolite subset. Metabolite C is directionally coupled with A and B in the sense that the presence of C in conserved pools forces A and B to be present in the same conserved pools and not vice versa (i.e., C is absent from pool 3, which encompasses A and B). Metabolite D is absent from all conserved pools and its concentration is not constrained by these pools.

FIGURE 6

Metabolic concentration coupling in glycolysis. Glycolysis admits three metabolite subsets, (G6P, F6P), (F1,6P, DHAP, G3P), and (3PG, 2PG). Directional coupling within glycolysis means that, for example, all conserved pools where PEP is present will always include both metabolite 1,3BPG and subset (3PG, 2PG).

FIGURE 7

Directional coupling between metabolite subsets for the genome-scale E. coli (iJR904) model.

FIGURE 8

Conserved pools spanning central metabolism of the genome-scale E. coli (iJR904) model. All metabolites within the four major central metabolism pathways (i.e., glycolysis, TCA cycle, PPP, and respiratory chain) belong to the four metabolite subsets, carbon influx/outflux, organic phosphate, energy and redox cofactors, and CoA subsets.