Precise metabolic engineering of carotenoid biosynthesis in Escherichia coli towards a low-cost biosensor

Micronutrient deficiencies, including zinc deficiency, are responsible for hundreds of thousands of deaths annually. A key obstacle to allocating scarce treatment resources is the ability to measure population blood micronutrient status inexpensively and quickly enough to identify those who most need treatment. This paper develops a metabolically engineered strain of Escherichia coli to produce different colored pigments (violacein, lycopene, and β-carotene) in response to different extracellular zinc levels, for eventual use in an inexpensive blood zinc diagnostic test. However, obtaining discrete color states in the carotenoid pathway required precise engineering of metabolism to prevent reaction at low zinc concentrations but allow complete reaction at higher concentrations, and all under the constraints of natural regulator limitations. Hence, the metabolic engineering challenge was not to improve titer, but to enable precise control of pathway state. A combination of gene dosage, post-transcriptional, and post-translational regulation was necessary to allow visible color change over physiologically relevant ranges representing a small fraction of the regulator's dynamic response range, with further tuning possible by modulation of precursor availability. As metabolic engineering expands its applications and develops more complex systems, tight control of system components will likely become increasingly necessary, and the approach presented here can be generalized to other natural sensing systems for precise control of pathway state.