Part:BBa_K2450401
OmpA SpyTag sfGFP label
OmpA with SpyTag Labelled with sfGFP
Usage and Biology
This part was designed to target proteins to the outer membrane and then outer membranes. As pathways targeting proteins to the outer membrane are not well understood, proteins must be bound to the outer membrane by other methods. The protein OmpA is abundant in the membrane, and efficiently targeted there by the Omp leader sequence.
The protein being targeted to the outer membrane is expressed as a fusion protein with spycatcher to bond it to OmpA in the outer membrane. The spytag/spycatcher system has been used to target proteins to the outer membrane by creating an isopeptide bond between the spytag and spycatcher domains. As spytag is very short it does not disrupt membrane proteins to the extent that a large fusion protein would, allowing it to fold properly in the membrane. This method has been shown to work by Alves et al in 'Protecting enzymatic function through directed packaging into bacterial outer membrane vesicles' 2016. For an example of a part designed to be targeted to the outer membrane in this way see part [BBa_K2450501].
sfGFP is used to show the protein has been successfully expressed and incorporated to the outer membrane. sfGFP is required as it is being targeted to the periplasm where GFP cannot fold.
Sequence and Features
- 10COMPATIBLE WITH RFC[10]
- 12COMPATIBLE WITH RFC[12]
- 21COMPATIBLE WITH RFC[21]
- 23COMPATIBLE WITH RFC[23]
- 25INCOMPATIBLE WITH RFC[25]Illegal NgoMIV site found at 4
- 1000COMPATIBLE WITH RFC[1000]
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