Part:BBa_K2220010

Secrete FliC-eGFP fusion protein

This part can secrete eGFP marked flagellin with the chasis S.cerevisiae.

α-Factor is a 13-residue peptide, secreted by cells of α mating type, that acts on cells of the opposite mating type to promote efficient conjugation between the two cell types, which leads to the formation of aα diploid cells. α-factor is synthesized as a precursor of 165 amino acids containing an 83-residue leader and four α-factor coding regions, each preceded by a short spacer peptide.As a natural signal peptide that is widely present in yeast, α-factor has strong and potent secretory properties. Studies have shown that its secretion is almost 100%, which can prevent the secretion of exogenous tubulin from interrupting the yeast's inherent microtubule system.

The flagellar filament is a helical structure composed of subunits of flagellin. And among various flagellin, the FilC has its own unique feature. The N- and C-terminal region of the FliC are important for filament formation, which are the D0 and D1 domain. While the D3 domain which is a central region of the flic playing no role in filament formation. Additionally, when FliC proteins are in the polymeric state, the D3 domain can be exposed on to the surface. So from above, we can replace the central region of flagellin by other proteins without destroying self-assembling ability and if we fuse the enzyme and FliC, we can obtain a filamentous material with high surface density of catalytic sites.

We use eGFP gene to replace the middle region of FliC gene,so that the eGFP can replace the D3 domain of flagellin. We add the alpha-factor to the N-terminal of the FliC-eGFP fusion protein. So it can secrete eGFP marked flagellin with the chasis S.cerevisiae.

Sequence and Features

Assembly Compatibility:

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

  Illegal NotI site found at 286
  
• 21
  INCOMPATIBLE WITH RFC[21]

  Illegal BamHI site found at 1785
  Illegal XhoI site found at 244
  
• 23
  COMPATIBLE WITH RFC[23]
• 25
  INCOMPATIBLE WITH RFC[25]

  Illegal AgeI site found at 595
  
• 1000
  COMPATIBLE WITH RFC[1000]