Part:BBa_K3013003

mamC-FbFP BS2 fusion

This part is designed as an improvement of part BBa_K1094401.

This fusion protein consists of mamC, a glycine linker and a fluorescent protein. MamC is suggested to be located in the magnetosome membrane, which are produced by magnetotactic bacteria. To report expression of mamC FbFP BS2 is used.

Magnetotactic bacteria produce magnetosomes under microaerobic conditions [Heyen U, Schüler D (2003) Growth and magnetosome formation by microaerophilic Magnetospirillum strains in an oxygen-controlled fermentor. Appl Microbiol Biotechnol 61:536–544]. As it requires elaborate equipment to setup such conditions, a simpler approach was tested by team Aachen 2019 which was cultivation of Magnetospirillum sp. and Rhodospirillum rubrum magneticum in closed flasks. Formation of magnetosomes was confirmed via transmission electron microscopy for R. rubrum magneticum and by formation of culture in a magnetic field for Magnetospirillum gryphiswaldense. Dissolved oxygen concentrations after days of cultivation tend towards zero. As maturation of GFP is dependent to oxygen, problems with reporting mamC expression levels under microaerobic or anaerobic conditions could arise [Tsien, R. Y. The green fluorescent protein. Annu. Rev. Biochem. 67, 509–544 (1998)]. Therefore, a more reliable reporter for microaerobic conditions with unknown oxygen concentrations is needed. Such a reporter is FbFP BS2 (BBa_K3013002). As its fluorophore formation is FMN-based (flavin mono nucleotide), it is not dependent to oxygen. We chose this fluorescent protein as it is also smaller than eGFP (eGFP: ~720 bp, FbFP BS2: 408 bp) and expression of the reporter should be as stressfree as possible for the host strain as well as the size of bigger fusion proteins like mamC-FbFP BS2-LCI (BBa_K3013000) should stay as small as possible for better expression results.

Usage and Biology

Sequence and Features

Assembly Compatibility:

• 10
  COMPATIBLE WITH RFC[10]
• 12
  COMPATIBLE WITH RFC[12]
• 21
  INCOMPATIBLE WITH RFC[21]

  Illegal BamHI site found at 498
  
• 23
  COMPATIBLE WITH RFC[23]
• 25
  INCOMPATIBLE WITH RFC[25]

  Illegal NgoMIV site found at 169
  Illegal NgoMIV site found at 253
  Illegal AgeI site found at 666
  
• 1000
  COMPATIBLE WITH RFC[1000]