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Project Description

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In the field of biodetection and bioremediation, despite numerous proofs of principle, most bioreporters and bioabsorbents have remained confined to the laboratory. Also bacterial reporters and absorbents have NOT been rational enough in design and NOT complex enough in function. Additionally, we notice that genetic engineering in this field needs streamline methods and it's time for a series of issues on the field application to be taken into consideration.

In this summer, we engineered our bacteria to resolve those hard truths mentioned above. MerR family transcription factors (TFs) was exploited to construct a series of biorepoters for heavy metal detection and bioabsorbents for heavy metal decontamination, based on Reverse Engineering principle.

MerR is a mercury responsive regulator. Primarily, we analyzed the function and operation of a MerR family TF, MerR in detail. Then modeling was conducted to analyze the characteristics of MerR and topology of its regulation behavior. Knowledge from the analyses enabled us to design genetic circuit that makes mercury sensor and regulator components more efficient and robuster. Our engineered bioreporters are able to discriminate concentration ranges of certain heavy metal in water independently of incubation time and cell activity in a wide window. This means that the field application can be carried out without the need for costly equipment while response quality and sensor sensitivity are still kept. Secondly, we constructed bioabsorbent strains that decontaminate mercury from polluted water. In our bioabsorbents, MerR was engineered into a single-chain, antiparallel, coiled coil peptide that mimics the metal binding domains, which reduces cost of metal binding. This was followed by inductive expression of engineered MerR protein on surface, periplasm and cytosol of E.coli cells. Mercury (II) absorption test showed that our bacteria can absorb more than 95% of 10^-7M Hg(II) in 5 minutes.

MerR family TFs share high homology at their metal binding domains, which implies that our strategies of bioreporter and bioabsorbent engineering might be applicable to other cases. We then expanded our reverse engineering strategy to another common toxic heavy metal, lead. A series of function tests showed that the lead bioreporters and bioabsorbents are comparable to mercury ones, proving validness of our engineering strategy. As almost each species of heavy metal has a corresponding MerR family TF, we can state that we have developed a streamlined method to construct heavy metal decontamination kits composed of valid bioreporters and bioabsorbents for field application.