Recent advances in cell-free synthetic biology have spurred the development of in vitro molecular diagnostics that serve as effective alternatives to whole-cell biosensors. However, cell-free sensors for detecting manmade organic water contaminants such as pesticides are sparse, partially because few characterized natural biological sensors can directly detect such pollutants. Here, we present a platform for the cell-free detection of one critical water contaminant, atrazine, by combining a previously characterized cyanuric acid biosensor with a reconstituted atrazine-to-cyanuric acid metabolic pathway composed of several protein-enriched bacterial extracts mixed in a one pot reaction. Our cell-free sensor detects atrazine within an hour of incubation at an activation ratio superior to previously reported whole-cell atrazine sensors. We also show that the response characteristics of the atrazine sensor can be tuned by manipulating the component ratios of the cell-free reaction mixture. Our approach of utilizing multiple metabolic steps, encoded in protein-enriched cell-free extracts, to convert a target of interest into a molecule that can be sensed by a transcription factor is modularly designed, which should enable this work to serve as an effective proof-of-concept for rapid field-deployable detection of complex organic water contaminants.

Last modified

• 02/19/2020

Creator

• Umut Akova
• Khalid K. Alam
• Michael C. Jewett
• Julius B. Lucks
• Adam D. Silverman

DOI

• https://doi.org/10.21985/n2-z5vp-tk94

Keyword

• biosensor
• TX-TL
• atrazine
• cell-free
• cyanuric acid
• metabolism
• synthetic biology
• transcription factor

Rights statement

• In Copyright