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    • December 7, 2018 at 13:31 #1997

      Eyad Fansa
      Participant

      Dear Colleagues,

      this is Eyad from the DPF facility in MPI-Dortmund. I was wondering whether someone have a good experience with the purification of human Caspase-1 from e.coli.

      I have the following three ideas:

      1- Separate expression of the p20 and p10 subunits followed by refolding from enclusion bodies.

      2- Repalcement of the autoproteolysis sites D119 and D279 by thrombine cleavage sites.

      3- Purify the full lenght protein in the prescence of Caspase inhibitor followed by inhibitor removal and autoactivation of the protein.

      I appreciate your comments or suggestions.

      Many Thanks,
      Eyad

    • December 7, 2018 at 13:52 #1998

      Anonymous
      Inactive

      Hi Eyad,

      did it before. Express full-length, refold and concentrate really, really high, this force autoactivation. After dilution you can polish with a final purification step.

      Cheers, Bernd

    • December 10, 2018 at 14:24 #1999

      Eyad Fansa
      Participant

      Hi Bernd,
      sounds good.
      Many Thanks, Eyad

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