Conclusion

Modified Pili Expression

Over the duration of the project we endeavored to answer a number of questions experimentally. Can we modify FimH with large proteins? Can the modified FimH be exported and can pilus biogenesis occur when large proteins have been fused? Can we modify the FimH protein at the N-terminal, after the signal peptide cleavage point? Can we drive expression of the fim operon with a constitutive promoter?

The T7_FimH_225_sfGFP construct exhibited fluorescence when examined using a plate reader (Tecan) and an Amnis ImageStream ISX. Protein expression was demonstrated using a Western blot and TEM-Immunogold techniques. The results in sum make a compelling case to suggest that a protein as large as sfGFP can be fused with FimH without losing the integrity of the tertiary structure or the robustness of pilus biogenesis.

Applied Design

Hydrocyclone

Metal Binding Reactor

Biosecurity

The results we obtained showed that a 10 minute exposure to UV was not able to cause sufficient cell death and therefore is not suitable for use as a biosecurity mechanism in our filtration system. In order to achieve the industry required escape frequency, which is of the order 10^-8, the water would have to be irradiated for a much more significant period of time which is not practical for our intended use. Therefore we need to explore possible alternatives, such copper alginate beads as detailed on the Applied Design page.