Imperial College London/M3/Detail

Module 3</b> is the final module of our system. It is designed so that the cell is programmed to destroy it's genetic material after encapsulation has finished.

After Module 2 has been completed, The E.ncapsulator is programmed to 'genetically self-destruct.' The cell produces enzymes which specifically target and cut DNA sequences, in order to destroy all genetic material contained within the cell. One advantage of using these enzymes for our 'killing' mechanism is that the cell membrane is left intact afterwards, and the protein of interest will still be protected by the encapsulated cell.

The E.ncapsulator requires the E. coli to be dead upon ingestion. This will prevent any transfer of genetic material between the bacterium and any gut microflora present, thereby avoiding any unexpected pathogenic effects. This is also especially important if the E.ncapsulator is to attain public acceptance, due to concerns over genetically modified organisms.

When

The thermally induced killing mechanism will only be triggered once encapsulation is complete.

How

Restriction enzymes found within bacteria and act as defense mechanisms against invading viruses. They work by recognising a certain DNA sequence of a few bases, and then cleaving the DNA strand. The E.ncapsulator is engineered to manufacture the restriction enzymes DpnII and TaqI when triggered, and these will cleave the genetic material within into fragments - thereby killing the cell.

As a protective mechanism against DNA destruction due to basal levels of restriction enzyme production, we have made use of the native E. coli Dam methylase protection system. This methylates DNA, which means that only high levels of restriction enzyme (ie. upon trigger) will cleave the DNA. is induced, rendering the bacterium no more than an inanimate shell containing our protein drug of choice.