Part:BBa_M50000

Streptavidin binding outer membrane protein

Assembly Compatibility:

• 10
  COMPATIBLE WITH RFC[10]
• 12
  COMPATIBLE WITH RFC[12]
• 21
  COMPATIBLE WITH RFC[21]
• 23
  COMPATIBLE WITH RFC[23]
• 25
  COMPATIBLE WITH RFC[25]
• 1000
  COMPATIBLE WITH RFC[1000]

Cell surface display protein with N-terminal and C-terminal streptavidin binding domain. eCPX-nSA-cNano was designed to be used to make E. coli bind to streptavidin coated magnetic beads, enabling bacterial capture. It however does not appear to work (see BBa_M50003).

Usage and Biology

Current bacterial display systems comprise of two parts: a scaffold protein that is anchored to the outer membrane, and a passenger peptide to be displayed^[1]. Here, we chose to display streptavidin-binding domains on the outer membrane of E. coli using Rice and Daugherty's engineered membrane protein eCPX as scaffold. eCPX is derived from the E. coli outer membrane protein OmpX. Both N and C terminus of eCPX are displayed on the extracellular side of the outer membrane, allowing for the possibility of biterminal fusion with streptavidin-binding domains, allowing for stronger ligand-substrate interactions and therefore more effective bacterial capture^[2]. Because an N-terminal fusion of eCPX with a 15 residue streptavidin-binding domain had already been created (eCPX-nSA), the streptavidin-binding peptide NanoTag was fused to the remaining free C-terminus of eCPX-nSA to form eCPX-nSA-cNano^[3].

References

[1] van Bloois, E.; Winter, R.T.; Kolmar, H.; Fraaije, M.W. 2011. Decorating microbes: surface display of proteins on Escherichia coli. Trends in Biotechnology. 29(2):79-86.

[2] Rice, J. J.; Daugherty, P. S. 2008. Directed evolution of a biterminal bacterial display scaffold enhances the display of diverse peptides. Protein Eng. Des. Sel. 21(7):435-442.

[3] Lamla, T.; Erdmann, V. A. 2004. The Nano-Tag, a streptavidin-binding peptide for the purification and detection of recombinant proteins. Protein Expression and Purification. 33:39-47.