Abstract
BackgroundE. coli can be used as bacterial cell factories for production of biofuels and other useful compounds. The efficient production of the desired products requires careful monitoring of growth conditions and the optimization of metabolic fluxes. To avoid nutrient depletion and maximize product yields we suggest using a natural mechanism for sensing nutrient limitation, related to biosynthesis of an intracellular messenger - guanosine tetraphosphate (ppGpp).ResultsWe propose a design for a biosensor, which monitors changes in the intracellular concentration of ppGpp by coupling it to a fluorescent output. We used mathematical modelling to analyse the intracellular dynamics of ppGpp, its fluorescent reporter, and cell growth in normal and fatty acid-producing E. coli lines. The model integrates existing mechanisms of ppGpp regulation and predicts the biosensor response to changes in nutrient state. In particular, the model predicts that excessive stimulation of fatty acid production depletes fatty acid intermediates, downregulates growth and increases the levels of ppGpp-related fluorescence.ConclusionsOur analysis demonstrates that the ppGpp sensor can be used for early detection of nutrient limitation during cell growth and for testing productivity of engineered lines.
Highlights
E. coli can be used as bacterial cell factories for production of biofuels and other useful compounds
To control the growth conditions during biotechnological applications of E. coli cells, we propose to use a biosensor, which couples changes in guanosine tetraphosphate (ppGpp) concentrations with a fluorescent Green fluorescent protein (GFP)-based output (Fig. 1)
The model describes the interrelationship between ppGpp, growth and Fatty acid (FA) production, and simulates the kinetics of GFP reporter of ppGpp, as summarised in the legend of Fig. 1 and discussed below
Summary
E. coli can be used as bacterial cell factories for production of biofuels and other useful compounds. Cells possess a natural mechanism of sensing of nutrient limitation related to the production of the second messenger guanosine tetraphosphate, ppGpp (an “alarmone”). This signalling pathway might be useful for the control of biotechnological processes. Regulation of the P1 and P2 promoters by ppGpp is fast (minutes) and covers a wide dynamic range of P1, P2 activities [6, 11] This suggests the possibility of transmitting changes in ppGpp concentrations into a P1, P2-coupled fluorescent output, which would allow continuous monitoring of transcriptional activity of ppGpp inside the cells. We explore the capacity of the biosensor to respond to dynamic changes in intracellular nutrient state during batch growth of fatty acid-producing and non-producing E. coli cells
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