Part:BBa_K4143339

TEV Protease + T4GALA Encapsulin Targeting Peptide

Adding this sequence to the end of a protein serves to target it for sequestration inside the T4GALA encapsulin (BBa_K4143337). This is because each cargo protein in an encapsulin can be identified by the presence of a short C-terminal targeting peptide at its end. [1][2]

Usage and Biology

For our project, we aimed to use the targeting peptide to facilitate the encapsulation of an antimicrobial peptide (AMP) inside an encapsulin. For this reason, we included a TEV protease site at the beginning of the targeting peptide for downstream AMP separation and isolation. The AMP sequence is described under part BBa_K4143336 and the encapsulin is characterized by part BBa_K4143337. To facilitate the encapsulation of our AMP, we attached the targeting peptide to the C-terminal end of the AMP.

AlphaFold Structural Characterization of AMP + Targeting Peptide

Because no structural information was available for our AMP + targeting peptide, we generated a tertiary structure using AlphaFold (Figure 1). Here, the targeting peptide is seen in the non-alpha-helical portion of the peptide, indicating it likely does not assume alpha helices or beta sheets for its secondary structure.

Figure 1: AlphaFold structural predictions for AMP HBCM2 + targeting peptide.

References

[1]T. W. Giessen and P. A. Silver, “Widespread distribution of encapsulin nanocompartments reveals functional diversity,” Nature Microbiology, vol. 2, no. 6, p. 17029, Mar. 2017, doi: 10.1038/nmicrobiol.2017.29.

[2]J. A. Jones, A. S. Cristie-David, M. P. Andreas, and T. W. Giessen, “Triggered reversible disassembly of an engineered protein nanocage,” bioRxiv, p. 2021.04.19.440480, Jan. 2021, doi: 10.1101/2021.04.19.440480.

Sequence and Features

Assembly Compatibility:

• 10
  COMPATIBLE WITH RFC[10]
• 12
  COMPATIBLE WITH RFC[12]
• 21
  COMPATIBLE WITH RFC[21]
• 23
  COMPATIBLE WITH RFC[23]
• 25
  COMPATIBLE WITH RFC[25]
• 1000
  COMPATIBLE WITH RFC[1000]