Difference between revisions of "Part:BBa K4035005:Design"
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===References=== | ===References=== | ||
(1) Chao, G.; Lau, W. L.; Hackel, B. J.; Sazinsky, S. L.; Lippow, S. M.; Wittrup, K. D. Isolating and Engineering Human Antibodies Using Yeast Surface Display. Nat Protoc 2006, 1 (2), 755–768. https://doi.org/10.1038/nprot.2006.94. | (1) Chao, G.; Lau, W. L.; Hackel, B. J.; Sazinsky, S. L.; Lippow, S. M.; Wittrup, K. D. Isolating and Engineering Human Antibodies Using Yeast Surface Display. Nat Protoc 2006, 1 (2), 755–768. https://doi.org/10.1038/nprot.2006.94. | ||
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| + | (2) https://www.uniprot.org/uniprot/P0CX80 | ||
Latest revision as of 20:14, 21 October 2021
Dimerization of the copper metallothionein 1 : CUP1-(EAAAK)3-CUP1
- 10COMPATIBLE WITH RFC[10]
- 12COMPATIBLE WITH RFC[12]
- 21COMPATIBLE WITH RFC[21]
- 23COMPATIBLE WITH RFC[23]
- 25COMPATIBLE WITH RFC[25]
- 1000INCOMPATIBLE WITH RFC[1000]Illegal SapI.rc site found at 384
Design Notes
The start and stop codon of the CUP1 genomic sequences were removed in order to fuse the proteins together. In addition, the full sequence was codon optimized before being ordered to avoid loops formation during synthesis. The synthetized CUP1-linker-CUP1 sequence was then inserted by Gibson Assembly in the pCTcon2-V5 plasmid (1) containing Aga2, V5 tag and the Gal1 promoter.
Source
The CUP1 sequence is the genomic sequence of the yeast copper metallothionein 1 protein (2). The sequence of the linker comes from a reverse translation of the amino acid sequence EAAAK and was codon optimized for the yeast S. cerevisiae. The fusion protein was fully synthetized.
References
(1) Chao, G.; Lau, W. L.; Hackel, B. J.; Sazinsky, S. L.; Lippow, S. M.; Wittrup, K. D. Isolating and Engineering Human Antibodies Using Yeast Surface Display. Nat Protoc 2006, 1 (2), 755–768. https://doi.org/10.1038/nprot.2006.94.