Part:BBa_K2292003

comDE two components signal transduction system

This is a composite used for producing comD, comE, two essential protein for the CSP two components signal transduction system. Originated from Streptococcus mutans. ComD is a kinase and membrane-associated receptor. After comD binded to the quorum sensing molecule CSP, the comD-CSP complex will phosphorylate comE. Then, the phosphorylated comE can turn on the transcription of downstream gene. FLAG-TAG is added after comE and c-Myc is added after comD

Figure 1 Mechanism of CSP detection

Figure 2 Agarose gel electrophoresis of PCR product (pSBBS1C)

Figure 3 Agarose gel electrophoresis of PCR product (pSBBS4S)

For the measurement of the fluorescent plate reader (excitation: 485 nm, emission: 520 nm), we found out that the best amount of bacteria for measuring, the CSP conc. is at the value of OD600 = 0.2 after a series of experiment.

Figure 4 Three-dimension diagram (Fluorescence-CSP concentration-Time)

The intensity of fluorescence slowly drops after a sharp rise in a small amount of time, hence we can tell that the reaction starts right away as soon as CSP is added.

Sequence and Features

Assembly Compatibility:

• 10
  COMPATIBLE WITH RFC[10]
• 12
  COMPATIBLE WITH RFC[12]
• 21
  INCOMPATIBLE WITH RFC[21]

  Illegal BglII site found at 268
  Illegal XhoI site found at 123
  Illegal XhoI site found at 153
  
• 23
  COMPATIBLE WITH RFC[23]
• 25
  COMPATIBLE WITH RFC[25]
• 1000
  COMPATIBLE WITH RFC[1000]