Coding

Part:BBa_K3582005

Designed by: Avadhoot Jadhav   Group: iGEM20_IISER-Pune-India   (2020-10-18)
Revision as of 10:52, 26 October 2020 by Avadhoot (Talk | contribs)


Inhibitory Sequence 1 Grafted In Cyclotide Kalata B1

This composite biobrick is believed to synthesize the readily deliverable cyclotide drug that could inhibit the 5LGD interaction between the host protein (CD36) and the parasite protein (CIDRa domain of PfEMP1). Different components involved in this biobrick are described below:

  • 1]BBa_K3582020: C intein is the modified version of the biobrick used by the Heidelberg 2014 team for circularization of the protein.
  • 2]BBa_K2333015: Bsa 1 Reversed recognition site
  • 3]BBa_K3582023(The cyclotide precursor): The gene coding for five loops of cyclotide Kalata B1 are inserted between the C Intein and the N intein so that it gets circularized after translation.
    • Strep II tag(Inbuilt): This tag is inserted in loop 3 of the cyclotide construct to reduce the haemolytic activity of the cyclotide and ease of purification.
  • 4]BBa_K3582012: Inhibitory peptide sequence 1 (IS1) grafted in loop 6 of the cyclotide structure to avoid the host-parasite protein interaction. This mutant sequence has a higher affinity for interaction as compared to wild type.
  • 5]BBa_K3582021: Bsa 1 recognition site
  • 6]BBa_K3582022: N intein is the modified version of the biobrick used by the Heidelberg 2014 team for circularization of the protein.

The inserted scars are basically the pieces of cyclotide precursor in whose loop the inhibitory peptide biobrick and strep II tag is grafted. The final translated protein product obtained using this biobrick is believed to act as a drug for preventing the CD36-PfEMP1 interaction.

The most interesting feature of this biobrick is that the inhibitory peptide sequence(BBa_K3582005) is replaceable which means other teams can replace that basic part with their own peptide sequences. In simple words, this biobricks provides a readily designed drug delivery system to the iGEM teams to insert their own inhibitory sequences of length 8-10 amino acids long to inhibit not just host-parasite protein interactions but whichever interaction they choose.


Figure 1. Structure of Kalata B1 grafted with strep II tag and inhibitor based on homology modelling.


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
    INCOMPATIBLE WITH RFC[25]
    Illegal AgeI site found at 112
  • 1000
    INCOMPATIBLE WITH RFC[1000]
    Illegal BsaI site found at 143
    Illegal BsaI.rc site found at 109

References

  • 1] The structural basis for CD36 binding by the malaria parasite.

Fu-Lien Hsieh1, Louise Turner2, Jani Reddy Bolla3, Carol V. Robinson3, Thomas Lavstsen2 & Matthew K. Higgins1

  • 2]Racemic and Quasi-Racemic X-ray Structures of Cyclic Disulfide-Rich

Peptide Drug Scaffolds** Conan K. Wang, Gordon J. King, Susan E. Northfield, Paola G. Ojeda, and David J. Craik*

Central (PMC). https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3330703/

  • 5] Poth, A. G. (2013, May 7). Cyclotides as Grafting Frameworks for Protein Engineering and Drug Design

Applications. PubMed. https://pubmed.ncbi.nlm.nih.gov/23893608/

  • 6] 2. Camarero, J. A. (2019, April 19). The Potential of the Cyclotide Scaffold for Drug Development.

MDPI. https://www.mdpi.com/2227-9059/7/2/31/htm#B31-biomedicines-07-00031


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