Difference between revisions of "Part:BBa K2450401"
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===Usage and Biology===
 
===Usage and Biology===
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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.
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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. Its usual function is as an ion channel in the outer membrane.  
  
 
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 [Alves et al 2016]. For an example of a part designed to be targeted to the outer membrane in this way see part [BBa_K2450501].
 
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 [Alves et al 2016]. For an example of a part designed to be targeted to the outer membrane in this way see part [BBa_K2450501].
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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.
 
 
    
 
    
  

Revision as of 12:03, 21 October 2017


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. Its usual function is as an ion channel in the outer membrane.

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 [Alves et al 2016]. For an example of a part designed to be targeted to the outer membrane in this way see part [BBa_K2450501].


Sequence and Features


Assembly Compatibility:
  • 10
    COMPATIBLE WITH RFC[10]
  • 12
    COMPATIBLE WITH RFC[12]
  • 21
    COMPATIBLE WITH RFC[21]
  • 23
    COMPATIBLE WITH RFC[23]
  • 25
    INCOMPATIBLE WITH RFC[25]
    Illegal NgoMIV site found at 4
  • 1000
    COMPATIBLE WITH RFC[1000]