Improved Parts
| Registry | Type | Name | Description |
|---|---|---|---|
| BBa_K4375007 | Coding | Codon optimized INPNC for surface display | INPNC (codes for N- and C- terminal domain of Ice Nucleation Protein (INP)) optimized for production in E. coli. |
| BBa_K4375018 | Device | MiniNano Display System | The MiniNano is a compact construct meant for EGFR detection via fluorescence. |
Parts Improvement
During our work, besides Nanobody Display System we developed another detection device, called MiniNano, which is a compact construct meant for extracellular protein (like tumor-specific receptors, antigens, etc.) detection via fluorescence.
Basically, we have broadened the functionality of an existing Part, INPNC (BBa_K811003), which was the work of Team UPenn from 2012. INPNC codes for N- and C- terminal domains of Ice Nucleation Protein (INP) from Pseudomonas syringe, and it is used for displaying proteins on bacteria's outer membrane. As a first step, we codon optimised it for protein expression in Escherichia coli (BBa_K4375007). After this, we linked INPNC to (by a flexible GS-linker, BBa_K4375010) a small, near-infrared emitting fluorescent protein, miRFP670 nano (BBa_K4375008), which is connected to an anti-EGFR specific Nanobody, 7D12, via llama-derived IgA hinge region linker (BBa_K4375011). Moreover, it contains a Myc epitope tag, so the specific interaction can be detected with labeled antibodies. This complex detection device is expressed under the constitutive promoter J23101* (BBa_K4375003) The Nanobody in the construct can be easily swapped with other protein domains, as its sequence is framed by BamHI and SphI restriction sites.
Although this construct may not fulfill the Part Improvement criteria because it lacks experimental results. Nevertheless, we thought that this Part should be mentioned on our Wiki, as it could be thought-provoking for other Teams in the years to come.