Part:BBa_K5466012

CadC intracellular domain

Intracellular domain of the EMeRALD sensing platform, consist of CadC ​​DNA ​​binding​ ​domain​ ​and ​​Juxtamembrane ​​region and an artificial Leu (16) transmembrane region. Upon dimerization active pCadBA (BBa_K3425101). Without stop codon for fusion with LBD for use in EMeRALD sensing platform.

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]

CadC

CadC is a one component system (OCS). OCS consist of single proteins containing both sensor and output domains. The most studied system is Escherichia coli’s CadC transmembrane transcriptional activator, on which modification relies the technology of EMeRALD (Engineered Modularized Receptor Activated via Ligand-induced Dimerization).

CadC is a bitopic transmembrane protein with a cytoplasmic split-DNA binding domain and a periplasmic pH-sensor in which dimerization of the sensor domain triggers dimerization of the cytoplasmic DBD and transcriptional activation.

EMeRALD

EMeRALD exploits the modular structure of this one-component system to generate new receptors by swapping sensor domains and introducing desired ligand binding domains (LBDs). This approach has been proven compatible with LBDs from other one-component systems as well as antibody-derived domains including VHH and scFvs, boosting the range of ligands of application.

You have to choose a ligand binding domain (LBD) to fuse with CadC, and choose a LBD distinct from the one used before, but that specifically binds to the molecule we intend for the receptor to detect to fuse with another CadC (unless the same LBD can dimerize in the presence of the ligand).

You can fuse it with a golden gate assembly without scars and in frame.

Reference

Chang, H., Mayonove, P., Zavala, A., De Visch, A., Minard, P., Cohen-Gonsaud, M., & Bonnet, J. (2017). A Modular Receptor Platform To Expand the Sensing Repertoire of Bacteria. ACS Synthetic Biology, 7(1), 166-175. https://doi.org/10.1021/acssynbio.7b00266

Chang, H., Zúñiga, A., Conejero, I., Voyvodic, P. L., Gracy, J., Fajardo-Ruiz, E., Cohen-Gonsaud, M., Cambray, G., Pageaux, G., Meszaros, M., Meunier, L., & Bonnet, J. (2021). Programmable receptors enable bacterial biosensors to detect pathological biomarkers in clinical samples. Nature Communications, 12(1). https://doi.org/10.1038/s41467-021-25538-y

Ulrich, L. E., Koonin, E. V., & Zhulin, I. B. (2004). One-component systems dominate signal transduction in prokaryotes. Trends In Microbiology, 13(2), 52-56. https://doi.org/10.1016/j.tim.2004.12.006