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Part:BBa_K4739004
Hcp-Minihepcidin fusion protein
The fusion protein was constructed based on BBa_K4739000(AAAGGG linker), BBa_K4739001(Minihepcidin), BBa_K4739002(Hcp), and a FLAG tag for antigen binding in western blot analysis. By replacing the sequence of Minihepcidin and FLAG tag with the sequence of required protein <25kD, the fusion protein should enable the T6SS of the engineered bacteria to deliver the protein into target or in its environment.
The T6SS needle's inner tube, vital for the functionality of T6SS, is constructed from hexamers of the Hcp protein, which are donut-shaped and stacked in a head-to-tail orientation to form a hollow tube. Its inner diameter is ∼40 Å and has an external diameter of ∼110 Å, limiting the size of effector <25kD (Cherrak et al., 2019). The Hcp hexameric rings and associated structures form the tube through which toxins or effectors can be transported during the secretion process. In our project, we utilize this structure to carry the drugs we aim to carry. For example, minihepcidin, and we witnessed a successful secretion through western blot. Results can be viewed from the result section of our wiki.
- 10COMPATIBLE WITH RFC[10]
- 12INCOMPATIBLE WITH RFC[12]Illegal NotI site found at 487
- 21COMPATIBLE WITH RFC[21]
- 23COMPATIBLE WITH RFC[23]
- 25INCOMPATIBLE WITH RFC[25]Illegal NgoMIV site found at 142
Illegal NgoMIV site found at 265 - 1000COMPATIBLE WITH RFC[1000]
Engineering and Results
Testing results can be viewed in BBa_K4739002.
Reference
Cherrak, Y., Flaugnatti, N., Durand, E., Journet, L., & Cascales, E. (2019). Structure and Activity of the Type VI Secretion System. Microbiology Spectrum, 7(4). https://doi.org/10.1128/microbiolspec.psib-0031-2019
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