Team:Imperial College London/Drylab/Protein Production

Protein Production

Based on the Genetic circuit, a LacI-IPTG inducible promoter is responsible for kickstarting the production of the drug.

• In the absence of IPTG, LacI represses the production of the drug (Cellulase or PAH)
• When IPTG is introduced, the LacI repressing pathway is “de-repressed”, and some output protein is produced.

Our goals

The modelling aims to provide an overview and better understanding of the M1 system’s function by:

• Characterizing the system.
• Modeling to account for several factors that may reduce/hinder the production of the protein drug such as:
  • Lac promoter leakiness
  • IPTG toxicity
  • Stability of output protein

This module is an integral part of the design, as large-scale commercialization of the drug of interest depends on finding the optimal conditions for protein production. We implemented a system of differential equations, having made some assumptions and predictions about how the system will behave.

  about the model assumptions and predictions!

The System

There are 6 differential equations that describe the behaviour of this system.
  about the equations and what they mean!

Summary of simulation results

• When we introduce IPTG into the system, it temporarily removes LacI from the system. Hence, during this period of time, we produce the drug of interest.
• When the effects of IPTG wear off, the system returns to equilibrium.
• The more IPTG we add in, the higher the amount of output protein.

  about the simulations!

• The effects of IPTG toxicity were investigated and we found that for these concentration ranges, IPTG is not toxic to cells. Click on the link to see the analyzed results: IPTG growth curves
• The constants in this model are arbitrary. We justify our usage of these values with a more detailed dynamical analysis of the system, which shows that it can only have fixed points[1]. System stability analysis (read about the equations before opening this file, it's intimidating!)

Conclusions

NOTE: These will be better understood once the reader has gone through the details ("Learn More").

• The greater the strength of the Lac promoter, the greater the repressive action of LacI prior IPTG induction.
• The greater the Lac promoter leakiness (k_leak) the greater the basal amount of expression of protein of interest, prior IPTG induction.
• The greater the amount of IPTG introduced, the greater the size of the bump in production of protein of interest.
• Here we assumed that the range of IPTG we have introduced is non-toxic for our cells. Growth curves will tell us whether IPTG does limit cell growth at the ranges we are interested in.

References

[1]Steven H. Strogatz (1994). Nonlinear dynamics and chaos: with applications to physics, biology chemistry and engineering. Addison Wesley. ISBN 0-201-54344-3
[2]3.Kuhlman T, Zhang Z, Saier MH Jr, & Hwa T (2007) Combinatorial transcriptional control of the lactose operon of Escherichia coli. - PNAS 104 (14) 6043-6048