M2 Genetic Circuit

Video

Genetic circuit

Elaboration

E.coli is naturally equipped with a multitude of defences to enable colonisation of the intestine. The E.ncapsulator is programmed to upregulate two global transcription factors (RcsB & YgiV) to hijack this natural process to maximise acid resitance. We have additionally upregulated a third enzyme (rfal) to enhance the encapsulation of single cells (over and above colony encapsulation). Finally, the two biosynthetic genes (OtsA & OtsB) code for the production of trehalose. Our manipulation of endogenous pathways reduces virulence while enchancing pill functionality.

This genetic circuit consists of two parts. The first part deals with the production of colanic acid, while the second part deals with the production of trehalose. Colanic acid is used to encapsulate the drug protein, protecting it from the harsh acidic environment of our gut. Trehalose will preserve the protein structure.

Genes RcsB, YgiV and Rfal control the production of colanic acid. RcsB is concerned with generating colanic acid, YgiV increases the yield and Rfal attaches the colanic acid produced, to the outer membrane of E. coli.

Genes OtsA and OtsB are involved in the production of trehalose.

Encapsulation is induced automatically when the concentration of glucose decreases below threshold. Initially, glucose concentration is high and this represses the CRP promoter. No transcription occurs and encapsulation does not occur. As E. coli grows, it consumes glucose, resulting in a decrease in glucose concentration until the threshold is reached. At this point, glucose will no longer be able to repress the CRP promoter. Colanic acid and trehalose would be produced.