Team:Manchester/Description1
Project Description
Introduction
The use of phosphate as a fertiliser is essential for maximizing crop plant yields in many agricultural settings. Phosphate runoff from fields can cause eutrophication. This leads to algal blooms and the deterioration of aquatic ecosystems. Moreover phosphate rock is a finite and increasingly scarce resource. Together, these two issues suggest a closed-loop solution. Our solution aims to address this issue through a system which could sequester and store high levels of phosphate: Phosphostore. Phosphostore uses genetically-modified bacteria that can accumulate phosphate from wastewater in protein-based microcompartments for future recycling. In our project, we expressed Eut (Ethanolamine utilisation) bacterial microcompartments and a polyphosphate kinase (PPK) enzyme with a Pdu (1,2-propanediol utilisation) localization tag. This enables the encapsulation of the PPK enzyme within the microcompartment, allowing the formation of polyphosphate chains to be built and stored safely from degradation by endogenous exopolyphosphatase (PPX). We were able to proof the concept of our project through fluorescent microscopy, which can be seen here.
Improving our System
We utilized a Design of Experiments approach to optimise the production of key components in our system.. Firstly, we explored the expression of of our key PPK enzyme. Through two rounds of DoE we were able to identify the optimal conditions for the expression of this enzyme, within the design space explored. DoE was also used to identify the optimum expression conditions for microcompartment expression. The five Eut proteins were split into 3 orthogonally controlled expression modules to allow us to tune the expression ratio of the microcompartment subunits.
An innovative ensemble modelling approach was also utilized to predict the operating characteristics of our phosphate starvation operon as a regulatory system for controlling microcompartment synthesis.
Real World
In interviews with a wide range of stakeholders, we explored the economic and regulatory constraints that would influence the implementation of our system in real-world water treatment plants, giving us detailed insights into the practices of innovation management in the water industry. We further investigated three key areas: intellectual property, GMO legislation and industrial scale-up which informed us to develop our extensive business plan. Investigation on intellectual property and GMO legislation in the UK inspired us our international collaboration with other iGEM teams to generate a worldwide legislation map, with the aim of assessing potential international markets where our project could be implemented. A continuous culture model was also incorporated in consideration of industrial scale-up to predict the rate at which Phosphostore devices could be produced on different substrates, allowing us to estimate the yearly cost of treating wastewater using Phosphostore.
