Part:BBa_K4375004

BLADE (light-responsive AraC transcription factor)

Figure 1: Schematic diagram showing the dimerization of the BLADE transcription factor.

BLADE is a modified AraC transcription factor engineered to react to blue light instead of arabinose. The AraC’s arabinose binding domain was swapped to a homodimerizing Vivid domain. Upon blue light illumination, the protein dimerizes and the AraBAD promoter becomes active.

Usage and Biology

Figure 2: Schematic diagram showing the working of the BLADE transcription factor.

BLADE (Blue light-inducible AraC dimers in Escherichia Coli) protein is engineered from the arabinose promoter (AraBAD) binding AraC protein by exchanging the dimerization domain of AraC with a Vivid domain (VVD), which homodimerizes upon light. The blue light sensing of VVD is because of FAD (flavin adenine dinucleotide) cofactor which changes conformation upon illumination and translates into the structure of the protein creating a new dimerization interface between the two domains. AraC is a monomer, and with this change, the heterodimerization changes to homodimerization which makes this system easier to control. Normally, one AraC monomer binds a high-affinity I1 site and another an O2 site on AraBAD, and when arabinose is added they dimerize and the O2 binding can now interact with a low-affinity I2 site which is needed for the RNA polymerase to bind the promoter. In BLADE arabinose is not needed and blue light illumination causes the VVD domains to dimerize and the AraC can bind to the I2 site. The wavelength requirement is 450 nm. The advantage of BLADE is that it is a photoinducible homodimerization system that requires only one gene instead of two.

Characteristics

For experimental results with this part please refer to BBa_K4375019 and BBa_K4375021.

Sequence and Features

Assembly Compatibility:

• 10
  COMPATIBLE WITH RFC[10]
• 12
  COMPATIBLE WITH RFC[12]
• 21
  COMPATIBLE WITH RFC[21]
• 23
  COMPATIBLE WITH RFC[23]
• 25
  COMPATIBLE WITH RFC[25]
• 1000
  COMPATIBLE WITH RFC[1000]

References

Romano, E.; Baumschlager, A.; Akmeriç, E. B.; Palanisamy, N.; Houmani, M.; Schmidt, G.; Öztürk, M. A.; Ernst, L.; Khammash, M.; Di Ventura, B. Engineering AraC to Make It Responsive to Light Instead of Arabinose. Nature Chemical Biology 2021, 17 (7), 817–827. https://doi.org/10.1038/s41589-021-00787-6.