Multi-Level Kinetic Model Explaining Diverse Roles of Isozymes in Prokaryotes

Abstract

Current standard methods for kinetic and genomic modeling cannot provide deep insight into metabolic regulation. Here, we developed and evaluated a multi-scale kinetic modeling approach applicable to any prokaryote. Specifically, we highlight the primary metabolism of the cyanobacterium Synechococcus elongatus PCC 7942. The model bridges metabolic data sets from cells grown at different CO2 conditions by integrating transcriptomic data and isozymes. Identification of the regulatory roles of isozymes allowed the calculation and explanation of the absolute metabolic concentration of 3-phosphoglycerate. To demonstrate that this method can characterize any isozyme, we determined the function of two glycolytic glyceraldehyde-3-phosphate dehydrogenases: one co-regulates high concentrations of the 3-phosphoglycerate, the other shifts the bifurcation point in hexose regulation, and both improve biomass production. Moreover, the regulatory roles of multiple phosphoglycolate phosphatases were defined for varying (non-steady) CO2 conditions, suggesting their protective role against toxic photorespiratory intermediates.

The Novel PII-Interacting Regulator PirC (Sll0944) Identifies 3-Phosphoglycerate Mutase (PGAM) as Central Control Point of Carbon Storage Metabolism in Cyanobacteria

“…Our results show that Sll0944 regulates the glycolytic carbon flux at the phosphoglycerate mutase step in a PII-dependent manner through interaction with 2,3-phosphoglycerate-independent phosphoglycerate mutase (PGAM) in response to the nitrogen status. This establishes PGAM was a key control point of cyanobacterial carbon flow, as predicted previously by kinetic modelling of the cyanobacterial low carbon response (20,21). Hence, we identified Sll0944 as PII-interacting protein that acts as a key regulator of PGAM in cyanobacterial carbon metabolism.…”

Different strategies of metabolic regulation in cyanobacteria: from transcriptional to biochemical control

“…As previously mentioned, Synechocystis 6803 cells did not show significant changes in the expression of genes for enzymes of central carbon metabolism triggered by changing CO 2 conditions 27 28 , whereas in Synechococcus 7942 those changes mainly explained the observed changes in the carbon metabolism in HC compared to AC cells 9 26 . It is notable that the mean value of transcriptional changes of the enzymes implemented into the model is 1.01 between HC and AC cells, whereas it was 1.36 in Synechococcus 7942.…”

“…We have extended the traditional kinetic model, based on metabolic data from one steady state, to multi-level kinetic model (various omics data and several steady states) and applied this approach to analyse the metabolic regulation of cyanobacterium Synechococcus 7942 14 26 . This multi-level kinetic modelling approach combines enzymatic data from the literature and experimentally obtained metabolic, fluxomic and transcriptomics data from several steady states to describe central carbon metabolism in this organism.…”

Identification of the light-independent phosphoserine pathway as an additional source of serine in the cyanobacterium Synechocystis sp. PCC 6803

“…The lack of mutants made it difficult to measure the relative importance of the two pathways. Thus, we tried to estimate it using an existing multi-scale kinetic model of Synechococcus that took into account existing metabolome and transcriptome data (Jablonsky et al, 2014). According to this model, we predict that in cells grown at 5 % CO 2 the phosphoserine pathway contributes to 66.4 % of serine synthesis, whereas due to the increased photorespiration it delivers only 28-37.6 % of serine under ambient CO 2 conditions (assuming 3-4.5 % oxygenation reaction of ribulose 1,5-bisphosphate carboxylase/oxygenase in cells grown at ambient air with 0.04 % CO 2 ).…”