Part:BBa_K1075025

pLac-RBS34-mCherry-(Ec)ssrA(DAS+4)-TT

The part contains the pLac promoter, a ribosomal binding site, the red fluorescent protein mCherry fused to the E. coli(ec)ssrA (DAS+4) tag and a double terminator.

biology

The (ec)ssrA tag is a short peptide sequence that can be fused to the C-terminus of proteins which should be degraded. The adaptor protein SspB binds to it and delivers the substrates to the protease ClpXP complex in E. coli. The mutation (DAS+4) in (ec)ssrA prevents the degradation of the ssrA-tagged protein without the presence of sspB and therefore makes the system more specific and controllable. [http://www.ncbi.nlm.nih.gov/pubmed/16762842]

mCherry is a red fluorescent protein with the excitation maximum at 587 nm and the Emission maximum at 610 nm. [http://www.ncbi.nlm.nih.gov/pubmed/15558047]

application

As we want to control protein degradation by controlling the function of ecSspB, we tagged the red fluorescent protein mCherry with ecssrA(DAS+4) to measure the degradation rate.

The part was designed for proof of principle. Application as a bacterial fotographic film might be possible as well.

Sequence and Features

Assembly Compatibility:

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

  Illegal NheI site found at 1717
  
• 21
  COMPATIBLE WITH RFC[21]
• 23
  COMPATIBLE WITH RFC[23]
• 25
  COMPATIBLE WITH RFC[25]
• 1000
  COMPATIBLE WITH RFC[1000]