Part:BBa_K2328008
usp45
Usage and Biology
We designed an expression and export system that enabled the targeting of the smurfp to specific locations in Lactococcus lactis cells, cytoplasm, cell wall, or medium. Optimization of protein secretion and of protein cell wall anchoring was performed with L. lactis cells by modifying the signals located at the N and C termini, respectively, of the reporter protein. Efficient translocation of precursor (∼95%) is obtained using the signal peptide from the lactococcal Usp45 protein and provided that the mature protein is fused to overall anionic amino acids at its N terminus; those residues prevented interactions of Nuc with the cell envelope. Nuc could be covalently anchored to the peptidoglycan by using the cell wall anchor motif of the Streptococcus pyogenes M6 protein. However, the anchoring step proved to not be totally efficient in L. lactis, as considerable amounts of protein remained membrane associated.
reference
[1] Dieye Y, et al. Design of a protein-targeting system for lactic acid bacteria. J Bacteriol. 2001.
Sequence and Features
- 10INCOMPATIBLE WITH RFC[10]Illegal PstI site found at 80
- 12INCOMPATIBLE WITH RFC[12]Illegal PstI site found at 80
- 21COMPATIBLE WITH RFC[21]
- 23INCOMPATIBLE WITH RFC[23]Illegal PstI site found at 80
- 25INCOMPATIBLE WITH RFC[25]Illegal PstI site found at 80
- 1000COMPATIBLE WITH RFC[1000]
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