Part:BBa_K2450451

OmpA SpyTag sfGFP tag

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].

For more information on how this part was used in our project, please see our wiki: http://2017.igem.org/Team:Oxford

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
  INCOMPATIBLE WITH RFC[1000]

  Illegal BsaI site found at 116