Coding

Part:BBa_K4244046

Designed by: Max Allewijn   Group: iGEM22_Wageningen_UR   (2022-10-08)
Revision as of 17:53, 8 October 2022 by MaxAllewijn (Talk | contribs)


flipUM (MMP-9 inducible)

"This flipUM is designed in such a way that it will only regain its colour when it is cleaved by MMP-9. Two inserts have been made, the E5 and K5 domain. These two domains dimerize and disrupt GFP structure. MMP-9 recognizes the cleavage site and cuts off the K5 domain, restoring Ultramarine function.

The flip design is based off the following paper: Zhang, Q., Schepis, A., Huang, H., Yang, J., Ma, W., Torra, J., Zhang, S. Q., Yang, L., Wu, H., Nonell, S., Dong, Z., Kornberg, T. B., Coughlin, S. R., & Shu, X. (2019). Designing a Green Fluorogenic Protease Reporter by Flipping a Beta Strand of GFP for Imaging Apoptosis in Animals. Journal of the American Chemical Society, 141(11), 4526–4530. https://doi.org/10.1021/JACS.8B13042/SUPPL_FILE/JA8B13042_SI_002.PDF"

The structure of flipUM was predicted using alphafold as shown below.

Flip ultramarine.png

The left shows uncleaved flipUM, the middle the cleaved, active flipUM, and the right regular UM

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 646
    Illegal XhoI site found at 703
    Illegal XhoI site found at 745
  • 23
    COMPATIBLE WITH RFC[23]
  • 25
    COMPATIBLE WITH RFC[25]
  • 1000
    COMPATIBLE WITH RFC[1000]


[edit]
Categories
Parameters
None