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Orthogonal Test in silico

Our Luminesensor is expected to be orthogonal to endogenous SOS pathway. In order to remove this obstacle on the application prospects of our Luminesensor, we used LexA408 instead of wild-type LexA DNA Binding domain. LexA408 and LexA are bio-orthogonal with each other since the sequence of the binding sites have variations (See Characterization).

Figure 10 <i>Luminesensor</i> LexA408-VVD

Figure 10. Moecular Modeling for Luminesensor LexA408-VVD.

By adding several nodes into the network, we constructed modeling for orthogonality in silica simulation:

Figure 11

Figure 11. Kinetic Network for Orthogonal Analysis


  • L denotes Luminesensor
  • I denotes the inner wild LexA
  • DL denotes the specific DNA binding site to Luminesensor
  • DI denotes the specific DNA binding site to wild LexA

The parameters are estimated as following:

k61.x10-4s-1dimered LexA releasing rate constant from non-specific binding site
K61.x10-2(n mol/L)-1dimered non-specific binding equilibrium constant

Table 2. Reaction Parameters for Orthogonality Simulation

Figure 12

Figure 12. Orthogonality Simulation Result.

The result shows that the contrast is highly related to the orthogonality. As our Luminesensor is orthogonal to the endogerous LexA system, our system still works well in bacteria with endogenously expressed LexA (See Characterization).


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