Part:BBa_K4810005
Modified M13 pVIII protein for Hippuric Acid binding (#2)
M13 pVIII protein is the major coat protein of the M13 bacteriophage, found in 2700 copies in the surface of the phage accounting for 98% of its mass. pVIII is used for peptide display for the discovery of peptides able to bind with specific ligands (biopanning). Modifications in the pVIII protein can be used to create M13 bacteriophage-based colorimetric biosensors for the detection of specific molecules. The sensor's colorimetric output is a result of structural changes caused by ligand-pVIII protein interactions. [1]
Coding sequence of the protein is modified to express the specific hexapeptide (ADIWRF) in the N-terminal of the protein, exhibiting high binding affinity with the volatile organic compound "Hippuric Acid", found in higher concentrations in the sebum of patients with Parkinson's disease.
The 23 amino acids of the N-terminal of pVIII protein undergo cleavage by a signal peptidase inside the bacterial cell, resulting in a functional protein of 50 amino acids. The specific modification occurred by deletion of the codons #2 to #5 of the functional protein and insertion of the respective codons of the desired peptide. Thus, the newly introduced peptide sequence is incorporated in the N-terminal of the protein.
The sequence of the specific peptide (ADIWRF) was determined by molecular docking simulations using Autodock Vina, opting for peptides with the highest affinity with Hippuric Acid and simultaneously the lowest affinity with other common organic compounds found in human sebum.
It should be noted that another part with the same function (BBa_K4810001) is also registered in iGEM's registry. Both sequences showed similarly satisfactory results by molecular docking simulations and thus both parts were designed, with the expectation of testing and comparing them.
References
[1]-Lee, J., Warner, C., Jin, HE. et al. Production of tunable nanomaterials using hierarchically assembled bacteriophages. Nat Protoc 12, 1999–2013 (2017). https://doi.org/10.1038/nprot.2017.085
Sequence and Features
- 10COMPATIBLE WITH RFC[10]
- 12COMPATIBLE WITH RFC[12]
- 21COMPATIBLE WITH RFC[21]
- 23COMPATIBLE WITH RFC[23]
- 25COMPATIBLE WITH RFC[25]
- 1000COMPATIBLE WITH RFC[1000]
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