Lux pL controlled luxR with lux pR controlled AHL degrading enzyme AiiA(lva tag).
This device has two protein generators: The first generator ‘luxpL-RBS-C0062-B0015’ produces an intermediate level of the LuxR protein.
The second generator ‘luxpR-RBS-C0060-B0015’ has a low basal expression of the AiiA enzyme. AiiA is capable of degrading the quorum sensing molecule AHL.
It should be noted that if AHL is added to this system it forms a complex with LuxR which will increase transcription of AiiA while reducing the transcription of LuxR.
Sequence and Features
- 10COMPATIBLE WITH RFC
- 12COMPATIBLE WITH RFC
- 21COMPATIBLE WITH RFC
- 23COMPATIBLE WITH RFC
- 25Illegal AgeI site found at 1283
- 1000Illegal BsaI.rc site found at 1101
Functionality of this part was tested by treating AHL mixtures with strains expressing this part. Liquid cultures were grown overnight, 1 mL aliquots of these cultures were spun down and resuspended in varying concentrations of AHL in PBS (to start the AHL degradation reaction). This was also done with a culture that contains BBa_K546002 (where aiiA is replaced with GFP).
After a 3h incubation, the supernatant was collected and subjected to a reporter strain: BBa_K546002. 200 µL samples of reporter strain/AHL mixtures were analyzed with a Molecular Devices Spectramax M2 spectrophotometer, detecting GFP. The result is shown below.
This part is functional, as AHL concentrations drop after incubation with E. coli transformed with this part. The GFP expression is much higher in the non treated mixtures, namely.
|control||(1) R0063 & (2) R0062|
|protein||(1) LuxR & (2) AiiA-LVA|