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Team:UCL London/Applications
From 2009.igem.org
Sponsors
Applications
The “Traffic-Light Stress Sensor” is intended to be used as biological, complementary tool to mechanical sensor during cell cultivation and bioprocessing. It is created to be sensitive to particular stresses within the E.coli cell such as misfolding of proteins and low Oxygen levels. The primary purpose for the application today is that it mainly should be used during process optimisation. The reason for this limitation is transcription and translation of fluorescent proteins is likely to have a negative metabolic impact on product titres during large scale commercial cell cultures.
Cell mediated detection of when pharmaceuticals proteins (produced in ‘’E.coli’’) are starting to fold improperly and aggregate could lead to much better understanding and design of bioprocesses and expression systems. Microorganisms like the enterobacterium Escherichia coli are great factories for recombinant expression of proteins. Another response encountered in recombinant systems is the accumulation of mis-folded protein in the cytoplasm. The aggregation of proteins in bacterial cells may have resulted from numerous factors such as:
- accumulation of high concentrations of folding intermediates
- inefficient processing by molecular chaperones
- changes in Temperature
Sufficient amount of oxygen transfer is a key parameter for maintaining high growth rate of ‘’E.coli’’. Additionally, during lower oxygen concentrations the ‘’E.coli’’ bacterium is re-adapting its metabolism to a state where it produces an increasing amount of acetate. Higher concentrations of acetate then become inhibitory for the cells. In large fermenters the oxygen level is measured via a DOT-meter. The data measurement device is fed-back, via a computer, to the motor of the impeller to increase agitation. Hence, the concentration of dissolved oxygen can be kept constant in response to the bacteria growing and consuming more oxygen. The DOT range, within which the process is being maintained, is often based on empirical experiments and experiences of what has worked on in the past. A biosensor that could signal rapidly from within the cells and give information about how they are adapting their metabolism to the present oxygen level; could shed light over obscure parameters in process design.
Moreover, within a larger fermenter are there also many so called “dead spots”. These spots are pockets where mixing is reduced. Usually, for the cells trapped in a dead spot, its effect will most rapidly be felt in the form of anoxia. Thus detecting low oxygen levels on an individual cellular level would enable determination of whether the dead spots are present in a reactor and this could possibly also facilitate the localization of dead spots.
