Part:BBa_K5466002:Design
AGA2P Signal Peptide
- 10COMPATIBLE WITH RFC[10]
- 12COMPATIBLE WITH RFC[12]
- 21COMPATIBLE WITH RFC[21]
- 23COMPATIBLE WITH RFC[23]
- 25COMPATIBLE WITH RFC[25]
- 1000COMPATIBLE WITH RFC[1000]
Design Notes
AG was added to the C-terminus based on findings from Wang et al. (2005). After cloning the scFv insert directly between the Aga2p leader sequence and the Aga2P protein, the fusion protein was not properly cleaved, resulting in no detectable signal. It was later discovered that, without the scFv insert, the EF dipeptide—derived from the EcoRI recognition site used to fuse the desired displayed protein with Aga2P—was positioned at the terminus of the Aga2 signal peptide. Previous studies indicated that the +1 and +2 amino acids at the N-terminus of the fusion protein are critical for signal peptidase processing. Several sequences (EF, EA, EAEA, and AG) were tested, all can be cut away by peptidase processing and successfully induced display. AG, being neutral and minimally bulky, was chosen as the default peptide spacer.
S. cerevisiae codon optimized.
Source
Wang, Z., Mathias, A., Stavrou, S., & Neville, D. M. (2005). A new yeast display vector permitting free scFv amino termini can augment ligand binding affinities. Protein Engineering Design And Selection, 18(7), 337-343. https://doi.org/10.1093/protein/gzi036