Part:BBa_K2649001

cpComE-NLS-VP16-NES SM6

This part contains a human optimized coding sequence for a constitutively phosphorylated ComE, the putative response regulator protein for the ComCDE two-component signaling pathway, native to Streptococcus mutans strain SM6 (a bacteria common in the human oral cavity). Design elements were chosen to optimize expression and function of ComE in human cells. The aspartic acid at amino acid position 60 (the putative phosphorylation site of the response regulator) in a wildtype ComE was changed to a glutamic acid, to act as a phosphomimetic. Normally, to be used as part of a functional sensing system, the ComE response regulator must be paired with the ComD histidine kinase receptor protein. When ComD binds to the S. mutans competence stimulating peptide, it autophosphorylates and transfers a phosphate group to an aspartic acid on ComE, enabling ComE to oligomerize and bind to specific DNA sequences to promote transcription. This constitutively phosphorylated ComE should theoretically always be able to bind DNA. The cpComE coding sequence is fused to a nuclear localization sequence (NLS), VP16 activating domain, and a nuclear export sequence (NES). It is in a pSB1C3 backbone and is BioBrick and MoClo compatible.

Sequence and Features

Assembly Compatibility:

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

  Illegal XhoI site found at 4
  
• 23
  COMPATIBLE WITH RFC[23]
• 25
  COMPATIBLE WITH RFC[25]
• 1000
  INCOMPATIBLE WITH RFC[1000]

  Illegal BsaI site found at 1