Modeling and experimental validation of the signal transduction via the Escherichia coli sucrose phospho transferase system

Abstract

Bacterial signal processing was investigated concerning the sucrose phosphotransferase system (sucrose PTS) in the bacterium Escherichia coli as an example. The about 20 different phosphotransferase systems (PTSs) of the cell fulfill besides the transport of various carbohydrates, also the function of one signal processing system. Extra- and intracellular signals are converted within the PTS protein chain to important regulatory signals affecting, e.g. carbon metabolism and chemotaxis. A detailed dynamical model of the sucrose PTS was developed describing transport and signal processing function. It was formulated using a detailed description of complex formation and phosphate transfer between the chain proteins. Model parameters were taken from literature or were identified with own experiments. Simulation studies together with experimental hints showed that the dynamic behavior of phosphate transfer in the PTS runs within 1 s. Therefore a description of steady state characteristics is sufficient for describing the signaling properties of the sucrose PTS. A steady state characteristic field describes the degree of phosphorylation of the PTS protein EIIA Crr as a function of the input variables extracellular sucrose concentration and intracellular phosphoenolpyruvate (PEP):pyruvate ratio. The model has been validated with different experiments performed in a CSTR using a sucrose positive E. coli W3110 derivative. A method for determining intracellular metabolite concentrations has been developed. A sample preparation technique using a boiling ethanol buffer solution was successfully applied. The PTS output signal degree of phosphorylation of EIIA Crr was also measured. Steady state conditions with varying dilution rate and dissolved oxygen concentration and dynamical variations applying different stimuli to the culture were considered. Pulse, and stop feeding experiments with limiting sucrose concentrations were performed. Simulation and experimental results matched well. The same holds for the expanded sucrose PTS and glycolysis model.