Part:BBa_K2830016
promoter+RBS+torA+GFP
TorA is a signal peptide that can transport proteins from gram-negative bacteria’s cytoplasm to periplasm, through twin arginine transport (TAT) pathway. TAT pathway depends on a tripartite complex of proteins, TatA, TatB plus TatC localized on the cytoplasmic membrane. the TatBC complex recognizes the signal sequence TorA and the folded protein is translocated via TatA into periplasm In SIAT-SCIE’s 2018 project, GFPmut3 is exported to periplasm to allow encapsulation by outer membrane vesicles (OMVs) that continuously bud off bacteria’s outer membrane. GFPmut3 serves to test the working efficiency of our TorA peptide by determining the ratio of the periplasmic fluorescence levels over the cytoplasmic fluorescence levels. Compared to other common transporting pathways that export proteins before folding (e.g. Sec and SRP-dependent pathways), TAT pathway is an available option for transportation of fully folded proteins. This is crucial since some GFP cannot fold properly in periplasm. Thus, they have to complete their folding in cytoplasm before they can be transported to periplasm. Consequently, TAT pathway is the only plausible option. TorA is not the only, but one of the most widely used signal peptide available for TAT pathway. Different papers report slightly different TorA sequences with varied length. Our sequence correspond to the TorA sequence of E.coli K-12 strain MG1655, which is chosen as it is the most common strain used in lab.
Sequence and Features
- 10COMPATIBLE WITH RFC[10]
- 12INCOMPATIBLE WITH RFC[12]Illegal NheI site found at 7
Illegal NheI site found at 30 - 21COMPATIBLE WITH RFC[21]
- 23COMPATIBLE WITH RFC[23]
- 25COMPATIBLE WITH RFC[25]
- 1000INCOMPATIBLE WITH RFC[1000]Illegal BsaI.rc site found at 822
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