title: New stable anchor protein and peptide linker suitable for successful spore surface display in B. subtilis.
abstract: BACKGROUND
  In last decade spores have been successfully used as a surface display platform. Various peptides or proteins were displayed this way as functional enzymes or antigens. Nearly all attempts involved use of three coat proteins: CotB, CotC or CotG. Increasing knowledge of the structure of the spore coat allowed us to propose the use of other proteins whose localization in the spore envelope has been determined. We also propose the application of a new linker suitable for building fusion proteins.
  
  
  RESULTS
  We show that a member of the outer coat, CotZ, is a good candidate as a new anchor protein useful in spore surface display. This protein allows use of relatively large passenger proteins and their efficient display on the spore surface. Analysis by Western- and dot-blotting, combined with immunofluorescence microscopy, allowed us to estimate the number of displayed fusion proteins molecules as 1.4 × 10(2) per spore. In addition, we present data indicating that the use of a peptide linker, which forms a stable α-helix, may greatly improve the display of anchored proteins on the spore surface.
  
  
  CONCLUSIONS
  CotZ can be used as an efficient anchor protein in the outer spore coat. Its localisation in the coat crust layer should guarantee surface display of passenger proteins. Moreover, a CotZ based fusion can tolerate relatively large passenger proteins for efficient spore surface display. In addition, to the properties of both the anchor and passenger proteins, an important issue is the nature of the linker. Here we present evidence that the linker, which forms a stable α-helix, may be crucial for successful display.
authors: Laboratory of Molecular Bacteriology, Intercollegiate Faculty of Biotechnology UG & MUG, Medical University of Gdańsk, Dębinki 1, Gdańsk 80-211, Poland.
authors: Krzysztof
authors: K
authors: Hinc
authors: Adam
authors: A
authors: Iwanicki
authors: Michał
authors: M
authors: Obuchowski
mesh_terms: D001412:Bacillus subtilis / Q000502:physiology*; D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism*; D005822:Genetic Vectors / Q000235:genetics / Q000378:metabolism; D008856:Microscopy, Fluorescence; D010455:Peptides / Q000235:genetics / Q000378:metabolism*; D011993:Recombinant Fusion Proteins / Q000032:analysis / Q000096:biosynthesis / Q000235:genetics; D013171:Spores, Bacterial / Q000378:metabolism*
chemical_list: D001426:Bacterial Proteins; D010455:Peptides; D011993:Recombinant Fusion Proteins