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===Reference===
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1) Shong, J., & Collins, C. H. (2013). Engineering the esaR promoter for tunable quorum sensing-dependent gene expression. ACS synthetic biology, 2(10), 568-575.
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Revision as of 12:44, 20 October 2021


Engineered quorum sensing esaR promoter: PesaR-C

A promoter that can interact with EsaR protein and inhibit downstream gene transcription.

Basic information

This part represents a promoter(Pesa) variant with two esa box, which can be bound to EsaR protein(see figure 2). esa box is a 18bps period located at a specific position of the promoter. After binding to the box, the EsaR will repress the recruitment of the RNA polymerase and inhibit the downstream transcription. However, when there are AHL molecules(such as 3OC6HSL) existing in the environment, they will interact with the EsaR and release it from the promoter, thus the transcription can be started.(Figure 1) The transcription level is controlled by the concentration of the AHL molecules.

Figure 1. Mechanism of the EsaR being an repressor under the control of the original PesaR. The red dots are the specific AHL molecule and in our project it is 3OC6HSL[1]
Below is the figure of the structure of the original promoter and the engineered one.
Figure 2. The structure of PesaR-C and the original PesaR.[2]
As we can see in figure 2, the original promoter: PesaR has only one esa box, which is at position -10. The PesaR-C has an additional esa box which locates between position -10 and -35. Previous studies has shown that the downstream gene expression level in this system depends on all these three factors: type of promoter, type of EsaR and concentration of the AHL molecule.(see figure 3)


21 UZurich PeaR charac.jpeg21 UZurich PeaRC charac.jpeg

Figure 3.Gene expression controlled by AHL molecules, EsaR regulator and PesaR promoter. On the left side, the test is based on the PesaR promoter and on the right side, the test is based on the PesaRC promoter. The luminescence level represents the gene expression level. It is clear that the regulation is controlled by all three factors which are mentioned above.

From figure 3 we can see that, the promoter PesaRC generally shows a higher transcription efficiency in comparison to PesaR. And the same EsaR regulator shows a lower sensitivity to the AHL molecule when it bind to PesaRC. When there are 10nM of AHL molecules in the environment, EsaR-D91G has already been saturated when it binds to PesaR but in PesaRC it doesn’t initiate the downstream gene expression yet.

Sequence and Features


Assembly Compatibility:
  • 10
    COMPATIBLE WITH RFC[10]
  • 12
    COMPATIBLE WITH RFC[12]
  • 21
    INCOMPATIBLE WITH RFC[21]
    Illegal BamHI site found at 281
    Illegal XhoI site found at 1
  • 23
    COMPATIBLE WITH RFC[23]
  • 25
    COMPATIBLE WITH RFC[25]
  • 1000
    COMPATIBLE WITH RFC[1000]


Reference

1) Shong, J., & Collins, C. H. (2013). Engineering the esaR promoter for tunable quorum sensing-dependent gene expression. ACS synthetic biology, 2(10), 568-575.