Part:BBa_K2871000
Map20, T3SS export signal peptide from Map gene
Map20 is a signal sequence encoding a 20 amino acid long signal peptide derived from the map gene from the Enteropathogenic E. coli strain E22. It is used as an N-terminal tag for a protein to be translocated by the E. coli type-3-secretion system (T3SS). This T3SS signal tag is described by Charpentier & Oswald (2004) to be sufficient to target proteins to E. coli type III secretion pathway. Its short length seemed to be attractive to us because we wanted to minimize folding interference between the signal sequence and the intended protein fused to the signal sequence.
Sequence and Features
- 10COMPATIBLE WITH RFC[10]
- 12COMPATIBLE WITH RFC[12]
- 21COMPATIBLE WITH RFC[21]
- 23COMPATIBLE WITH RFC[23]
- 25INCOMPATIBLE WITH RFC[25]Illegal AgeI site found at 85
- 1000COMPATIBLE WITH RFC[1000]
Experimental characterization
Map20-fusion protein expression plasmid construction
In order to be able to test the expression of N-terminal Map20-fusion protein, we constructed a plasmid consisting of a Map20-tagged reporter gene controlled by pBAD promoter, which can be activated by adding arabinose to the culture media. The Map20 T3SS signal is linked with the reporter protein by glycine-serine linker peptide to decrease folding interference. 6X-His tag has been added to the C-terminal to serve as an epitope tag for western blot. The plasmid map is shown in Figure 1.
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