Difference between revisions of "Part:BBa K3117046"
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===Usage and Biology=== | ===Usage and Biology=== | ||
| − | By connecting the variable regions of the heavy and light chain of an anti-GPA33 antibody with a short, flexible GGGGS linker (<partinfo>BBa_K3117028</partinfo>), the scFv retains it's antigen-binding ability and is much smaller than a conventional antibody. The SpyCatcher attached to the scFv belongs to the SpyTag/SpyCatcher system, of which the sequence is derived of the FbaB protein in the bacteria Streptococcus pyogenes. Once it comes into contact with its corresponding other part, the SpyTag (<partinfo>BBa_K3117037</partinfo>), they bind covalently (Hatlem | + | By connecting the variable regions of the heavy and light chain of an anti-GPA33 antibody with a short, flexible GGGGS linker (<partinfo>BBa_K3117028</partinfo>), the scFv retains it's antigen-binding ability and is much smaller than a conventional antibody. The SpyCatcher attached to the scFv belongs to the SpyTag/SpyCatcher system, of which the sequence is derived of the FbaB protein in the bacteria Streptococcus pyogenes. Once it comes into contact with its corresponding other part, the SpyTag (<partinfo>BBa_K3117037</partinfo>), they bind covalently (Hatlem et al., 2019). This allows our part to be used in a modular manner in combination with other molecules carrying the SpyTag. The sequence contains a C-terminal His-Tag (<partinfo>BBa_K3117005</partinfo>) for easy purification and detection. Secretion of the protein into the perisplasm is ensured by the pelB sequence leader (<partinfo>BBa_K3117012</partinfo>). When the protein passes the membrane, the leader segment is cleaved off. |
Since the scFv targets GPA33, which is expressed on more than 95% of colon cancers (Rageul et al., 2009), our part is expected to be optimal for specifically directing its fusion partner (e.g. an effector protein) to colon cancer cells. | Since the scFv targets GPA33, which is expressed on more than 95% of colon cancers (Rageul et al., 2009), our part is expected to be optimal for specifically directing its fusion partner (e.g. an effector protein) to colon cancer cells. | ||
Revision as of 21:12, 21 October 2019
scFv bispecific antibody against GPA33 and CD3 codon optimized for E.coli
The part BBa_K3117026 is a fusion protein of an anti-GPA33 single-chain variable fragment (scFv) and SpyCatcher (BBa_K3117016). This part is codon optimized for the expression in E.coli.
Usage and Biology
By connecting the variable regions of the heavy and light chain of an anti-GPA33 antibody with a short, flexible GGGGS linker (BBa_K3117028), the scFv retains it's antigen-binding ability and is much smaller than a conventional antibody. The SpyCatcher attached to the scFv belongs to the SpyTag/SpyCatcher system, of which the sequence is derived of the FbaB protein in the bacteria Streptococcus pyogenes. Once it comes into contact with its corresponding other part, the SpyTag (BBa_K3117037), they bind covalently (Hatlem et al., 2019). This allows our part to be used in a modular manner in combination with other molecules carrying the SpyTag. The sequence contains a C-terminal His-Tag (BBa_K3117005) for easy purification and detection. Secretion of the protein into the perisplasm is ensured by the pelB sequence leader (BBa_K3117012). When the protein passes the membrane, the leader segment is cleaved off.
Since the scFv targets GPA33, which is expressed on more than 95% of colon cancers (Rageul et al., 2009), our part is expected to be optimal for specifically directing its fusion partner (e.g. an effector protein) to colon cancer cells.
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]
References
1. Hatlem, D., Trunk, T., Linke, D., & Leo, J. C. (2019). Catching a SPY: Using the SpyCatcher-SpyTag and Related Systems for Labeling and Localizing Bacterial Proteins. International journal of molecular sciences, 20(9), 2129.
2. Rageul, J., Mottier, S., Jarry, A., Shah, Y., Théoleyre, S., Masson, D., . . . Denis, M. G. (2009). KLF4‐dependent, PPARγ‐induced expression of GPA33 in colon cancer cell lines. International journal of cancer, 125(12), 2802-2809.
3. Schoene, C., Fierer, J. O., Bennett, S. P., & Howarth, M. (2014). SpyTag/SpyCatcher cyclization confers resilience to boiling on a mesophilic enzyme. Angewandte Chemie International Edition, 53(24), 6101-6104.