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Revision as of 22:49, 11 October 2022


Trim-(GSGS)-Coh2

This parts code for the trim21 E3 ligase having his PRYSPRY domain truncated (BBa_K3396007), fused to type 1 Cohesin module derived from Clostridium thermocellum cellulosome scaffoldin using Glycine serine flexible linker repeated two times to maintain part flexibility needed during target ubiquitination

Usage and Biology

this part is the main part in our Snitch system, it is supposed to bind to the protac which in turn will bind to tau protein. The hypothesis is when the binding occurs between the three parts, Trim21 will recruit E2 conjugating enzyme that carries ubiquitin and move the ubiquitin from the E2 to tau. After ubiquitination, tau protein is supposed to be degraded by 26S proteasome

Sequence and Features

the sequence was optimized for E.coli expression


Assembly Compatibility:
  • 10
    COMPATIBLE WITH RFC[10]
  • 12
    INCOMPATIBLE WITH RFC[12]
    Illegal NheI site found at 225
  • 21
    COMPATIBLE WITH RFC[21]
  • 23
    COMPATIBLE WITH RFC[23]
  • 25
    INCOMPATIBLE WITH RFC[25]
    Illegal NgoMIV site found at 182
  • 1000
    COMPATIBLE WITH RFC[1000]

Modeling

tTrim21-GSGS-Coh2 is modeled by AlphaFold2, ITASSER, Robetta and TrRosetta, best model obtained from TrRosetta. 

                             Figure 1.: Predicted 3D structure of our fusion protein tTrim21-GSGS-Coh2.


Table 1: Quality assessment parameters of tTrim21-GSGS-Coh2. model.



References

1- Carvalho AL, Dias FM, Nagy T, Prates JA, Proctor MR, Smith N, Bayer EA, Davies GJ, Ferreira LM, Romão MJ, Fontes CM, Gilbert HJ. Evidence for a dual binding mode of dockerin modules to cohesins. Proc Natl Acad Sci U S A. 2007 Feb 27;104(9):3089-94. doi: 10.1073/pnas.0611173104. Epub 2007 Feb 20. PMID: 17360613; PMCID: PMC1805526.