Team:Fudan-CHINA/Basic Part
Best basic part: BBa_K2886002
We present BBa_K2886002 as our best basic part which encodes VEGF-scFv that can recognize VEGF.
VEGF (vascular endothelial growth factor), originally known as VPF (vascular permeability factor), is a ligand commonly used in research. As a kind of endogenous growth factor produced by both human and mouse cells, VEGF has various biological effects such as stimulating the formation of blood vessel and enhancing the permeability of it. VEGF is a big family, consisting of VEGF-A, VEGF-B, VEGF-C, VEGF-D, VEGF-E and PGF, of which VEGF-121 and VEGF-165 are two main types in body as well as used in research.
VEGF-scFv, standing for VEGF single-chain variable fragment, is adapted from the outermost Ig-like domain of the antibody of VEGF. VEGF-scFv encoded by BBa_K2886002 is a fusion protein containing light chain and heavy chain linked by a GS linker. As for its function, VEGF-scFv can bind many kinds of VEGF including VEGF-121 and VEGF-165, the most commonly used subtypes of VEGF referred above. Thus, iGEMers could use this part to achieve the detection of most kinds of VEGF efficiently and conveniently with our part.
VEGF (vascular endothelial growth factor), originally known as VPF (vascular permeability factor), is a ligand commonly used in research. As a kind of endogenous growth factor produced by both human and mouse cells, VEGF has various biological effects such as stimulating the formation of blood vessel and enhancing the permeability of it. VEGF is a big family, consisting of VEGF-A, VEGF-B, VEGF-C, VEGF-D, VEGF-E and PGF, of which VEGF-121 and VEGF-165 are two main types in body as well as used in research.
VEGF-scFv, standing for VEGF single-chain variable fragment, is adapted from the outermost Ig-like domain of the antibody of VEGF. VEGF-scFv encoded by BBa_K2886002 is a fusion protein containing light chain and heavy chain linked by a GS linker. As for its function, VEGF-scFv can bind many kinds of VEGF including VEGF-121 and VEGF-165, the most commonly used subtypes of VEGF referred above. Thus, iGEMers could use this part to achieve the detection of most kinds of VEGF efficiently and conveniently with our part.
Moreover, to realize higher affinity and efficiency of ligand-receptor binding, we alter a few amino acids on the binding surface with the assist of Rosetta.
Actually, we are not the first one to introduce VEGF-scFv to iGEM. Team NCTU-Formosa did this work in the year 2015 (BBa_K1694003). Nevertheless, to our regret, we find that there is no experimental structure in PDB (Protein Data Base), which may decrease the reliability of the outcome if we optimize VEGF-scFv based on this part. Thus, this year we introduced a new type of VEGF-scFv (BBa_K2886002) that has accepted structure in PDB, enabling us to work out a much more credible improvement. And, we also simulate the structure of BBa_K1694003 to help us learn about our improvements.
As is vividly depicted in the Figure 1, we can find that compared to the binding surface of their original version of BBa_K2886002, the mutant version has a lower interface score, which indicates that our work does make sense. For more detailed information of this process, please click here to visit our Receptor Optimization page.
Actually, we are not the first one to introduce VEGF-scFv to iGEM. Team NCTU-Formosa did this work in the year 2015 (BBa_K1694003). Nevertheless, to our regret, we find that there is no experimental structure in PDB (Protein Data Base), which may decrease the reliability of the outcome if we optimize VEGF-scFv based on this part. Thus, this year we introduced a new type of VEGF-scFv (BBa_K2886002) that has accepted structure in PDB, enabling us to work out a much more credible improvement. And, we also simulate the structure of BBa_K1694003 to help us learn about our improvements.
Figure 1. The structure of VEGF-scFv and the interface score of modified version of BBa_K2886002 compared with original version and BBa_K1694003. (a) VEGF-scFv is two outermost Ig-like domains of the antibody of VEGF joined together with a GS linker. (b) According to the interface scores which indicate the affinity of VEGF-scFv to VEGF of revised BBa_K2886002, original BBa_K2886002 and BBa_K1694003, we can draw the conclusion that revised BBa_K2886002 probably has a higher affinity to VEGF after our modification.
What we do has set a good example for iGEM teams. Using similar tool and strategy, you can make improvements to your target protein to make it more practical or we can even endow new functions to a protein theoretically, which is definitely promising in the field of synthetic biology.
[1]Lai, Y. et al., Serum VEGF levels in the early diagnosis and severity assessment of non-small cell lung cancer. J CANCER 9 1538 (2018).
[2]Wikipedia contributors. "Vascular endothelial growth factor." Wikipedia, The Free Encyclopedia. Wikipedia, The Free Encyclopedia, 27 Aug. 2018. Web. 17 Oct. 2018.
Address
G604, School of Life Sciences, Fudan University
2005 Songhu Road, Yangpu, Shanghai, China
2005 Songhu Road, Yangpu, Shanghai, China
