R-BphP

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Revision as of 09:54, 9 June 2009

The BphP / PpsR system

PpsR1 is a redox sensitive activator. It binds DNA under anaerobic conditions, and forms a tetramer via a disulfide bond. This interaction is ablated in the mutant PpsR1-C429S; meaning that we should be able to mimic an anaerobic system even under aerobic conditions.

PpsR2 is a transcriptional repressor, regulated by BphP.

PpsR1 and PpsR2 bind to TGTN_12ACA possibly arranged in tandem with a 7 base spacer. The affinity for both PpsRs are around 100nM.

BphP (or BrBphP for the Bradyrhizobium variant) is sensitive to far-red light (~770nm) and controls PpsR1.

To find

- Define genetic circuit Tú
- Look for exact wavelength for the corresponding light receptor Basile
- Sequences of the genes involved in the pathway (minimal genes to => Pubmed) Tú

Implementation

There are two modes in which we can use the system. The minimal complement requires PpsR2 and its regulator BphP. We can generate a hybrid system that uses a well known activator or transcription or a constitutively active promoter which will be repressed via PpsR2 upon exposure to far-red light (770nm).

It is also possible to reconstitute the entire system including PpsR1-C429S, which will serve as the activator and will be de-coupled from the oxidative state of the cell due to the ablation of the disulfide bond formation.

To do

contact Eric Giraud (giraud@mpl.ird.fr) and ask if he could send us the following material:
- Bradyrhizobium ORS278
- The following plasmids:
  - pBAD::ppsR1
  - pBAD::ppsR2
  - pGEM-T::ppsR2/BrbphP
- the PpsR1-C429S construct

Found

- Bacterium: Bradyrhizobium can be ordered on ATCC

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@@ Line 7: / Line 7: @@
 PpsR1 and PpsR2 bind to TGTN_12ACA possibly arranged in tandem with a 7 base spacer. The affinity for both PpsRs are around 100nM.
-BphP (or BrBphP for the Brayrhizobium variant) is sensitive to far-red light (~770nm) and controls PpsR1.
+BphP (or BrBphP for the Bradyrhizobium variant) is sensitive to far-red light (~770nm) and controls PpsR1.
 ==To find==