Central Carbon Metabolism of Sulfolobus solfataricus

Silicon cell model for the central carbohydrate metabolism of the archaeon Sulfolobus solfataricus under temperature variation

By varying the input or output flux via PGK and FBPAase manipulation respectively the pathway efficiency can be optimised.

Mathematical model of a subset of reactions comprising the three most temperature sensitive intermediates of the gluconeogenic pathway in S. solfataricus

Experimental analysis of pathway fluxes with varying PGK and FBPAase concentrations in the incubation assay.

Varying the relative amounts of PGK and FBPAase and analyse the 3PG uptake and F6P production rates.

Mathematical model for the reconstituted system with PGK, GAPDH, TPI and FBPAase.

The four purified enzymes were incubated in assay buffer and consumption of 3PG and production of F6P were measured in time, together with GAP and DHAP concentrations.

Mathematical model for the analysis of carbon loss of BPG, GAP and DHAP in reconstituted system of S. solfataricus at 70C.

Model simulation showing the effect of varying PGK or FBPAase concentration on the efficiency for 3PG to F6P conversion.

Combined plot fo the model simulations and the experimental data for the PGK and FBPAase titrations.

Increasing the amount of PGK in the incubation assay and measuring the 3PG and F6P fluxes.

A time simulation of the mathematical model is shown together with the experimental data to which the model was fitted.

A time simulation of the mathematical model is shown together with the experimental data to which the model was fitted.

Mathematical model for TIM kinetics, GAP and DHAP saturation and PEP inhibition.

Mathematical model for GAPDH kinetics, saturation for GAP, BPG, NADP, NADPH, Pi.

Mathematical model for FBPAase kinetics

An exponential decay model was fitted to the experimental data set of DHAP degradation at 70C. The model is uploaded as an SBML file. A data file showing the model fit to the experimental data set is included. The rate constant for DHAP degradation at 70C was estimated to be 0.0225 1/min, equivalent to a half life time of 30.8 min.

An exponential decay model was fitted to the experimental data set of GAP degradation at 70C. The model is uploaded as an SBML file. A data file showing the model fit to the experimental data set is included. The rate constant for GAP degradation at 70C was estimated to be 0.056 1/min, equivalent to a half life time of 12.4 min.

The recombinant enzymes were purified from the cell free extracts of E. coli expressing the respective enzymes.

Preparation of cell free extracts of the recombinant E. coli strains expressing the respective S. solfataricus enzymes.

Cloning and heterologous expression of gluconeogenic enzymes in E. coli