Part:BBa_K365013

ClpX trimer

We fused 3 ∆N-ClpX with 20 aa linker in order to increase the stability of ClpX pseudo-hexamer.

Background:

Phytochrome B needs to be fixed to the Clpx hexametric part of the protease. It is possible to link one phytochrome B per ClpX monomer but this could lead to steric problems. So the decision was made to follow the idea of the publication of Tanja Baker 2009. This permits to have only one phytochrome B for three ClpX units. Moreover the publication proved that the speed of the assembly of two ClpX trimers is quite the same than with ClpX monomers. So the Clpx hexameric part is composed of two ClpX trimers each one couple with a phytochrome B.


You can visit our wiki "visual description" page for more info

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 699
  Illegal BglII site found at 1863
  Illegal BglII site found at 3027
  
• 23
  COMPATIBLE WITH RFC[23]
• 25
  COMPATIBLE WITH RFC[25]
• 1000
  COMPATIBLE WITH RFC[1000]

Conception: ClpX primer was constructed due to cloning methods. The new approach in this method is that the primer was built from a PCR product. The construction of the ClpX dimer was conducted with standard methods. Then in a PCR reaction the ClpX primers located inside of the dimer were used to construct this new primer. The product of this PCR was used as primer for constructing the ClpX trimer.