Team:SUIS Shanghai/Model
Model
In order to ensure we would be able to detect vibrioferrin production in our culture broth after induction we sought to gain insight from previous iGEM models, that would allow us to estimate how long after induction with IPTG would our cells be producing and secreting vibrioferrin at a constant rate. We used information from a previous model by the Evry 2013 iGEM team, who, as part of their project “Iron Coli”, also worked with siderophores and E.coli (although they were measuring enterobactin). We found that components of their model may be useful to our project and our question because of the mechanisms of both siderophores production. The biosynthesis of both siderophores requires the stepwise reactions of enzyme catalysis. We felt our project could gain insight from this model as it considers the delay in enzyme production as well as the overall siderophore production time of four reactions, similar to the production of vibrioferrin by our gene cluster.
https://2013.igem.org/Team:Evry/Modelmeta3
We developed our own equations for the production of vibrioferrin. However we had no kinetic parameters available to use the equations as a predictive tool. It was decided that at best, we could use the previous model as a guide and to provide an estimate of the vibrioferrin production time. This model also produced a siderophore via a gene cluster of similar length in e.coli, however their system was under control of the Ferric uptake regulator (Fur box). Despite this we gained insight into an estimation of how long our system could produce vibrioferrin at a constant rate.
From this insight we decided to incubate for at least 2 hours after induction of our T7 promoter before centrifuging to create a cell free supernatant containing vibrioferrin.
Where the velocities of the reactions are given by:
Our equations can be used to model vibrioferrin production, after T7 promoter induction, in our system if appropriate Km and Kcat values are learned from experiment.
Figure 1 - Model predicting enterobactin production in e.coli is a much slower process than the production of enzymes. This model predicted approximately a 2 hour time period for the production of enterobactin at a constant rate. We used this model to provide a rough prediction of how long our engineered cell would produce vibrioferrin at a constant rate.
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