Team:Groningen/Brainstorm/Glucose Sensing

From 2009.igem.org

Originally the Rainbow bacteria, we now no longer plan to use the Cre/Lox system, but rather focus on glucose sensing. This choice was made because we have no good way to control the cre/lox recombination the way we would need for glucose sensing.

The idea now is to create a bacterium that senses the outside glucose concentration and produces an inversely proportional response.

Requirements

We use the [http://en.wikipedia.org/wiki/MoSCoW_Method MoSCoW prioritization method]:

• Must have this:
  • Response should be inversely proportional to the outside glucose concentration.
  • It should be measurable on-line (visually for example).
• Should have this if at all possible:
  • Response should be a gradient (not just on/off).
  • Delay in the response should not exceed roughly 5-10 minutes.
  • (Model) parameters should be known or obtainable.
• Could have this if it does not affect anything else:
  • Response should reasonably accurately follow the concentration (after a delay).
• Won't have this now but would like in the future:
  • Expand to other metabolisms.
  • Produce something more useful than a colour.

Design

Our bacterium would have the following components:

• [http://www.genome.jp/dbget-bin/www_bget?pathway:ko02060 The PTS system], which is already present in ... Which organisms?
• A promoter that hooks into the PTS system. Can be either:
  • Constitutively active, but repressed by CcpA.
  • Activated by CcpA.
• Product behind promoter as response.
• Degradation-tag attached to the product to increase turnover rate.

Modelling

First of all we are trying to model the PTS system as is. We're looking at the following existing models:

• A limited model consisting of only 9 substances and 7 reactions, as described in Neves1999.
• A more complete model [http://jjj.biochem.sun.ac.za/cgi-bin/processModelSelection.py?organism=Lactococcus+lactis&category=metabolism by M.H.N. Hoefnagel, J. Hugenholtz and J.L. Snoep] (hoefnagel2), discussed in Hoefnagel2002.

Our attempts at using these models are available in [http://igemgroningen.googlecode.com/svn/trunk/glucose our SVN repository].

Once we have a model of the PTS system we will try adding one or more "reactions" for the HPr phosphorylation/dephosphorylation under the influence of FBP and ATP, as this leads to HPr-ser46, which lets Ccpa bind to a cre site more easily. For the moment it is assumed that HPr and Ccpa are available in abundance.