A recombinant bacterial cell surface (S-layer)-major birch pollen allergen-fusion protein (rSbsC/Bet v1) maintains the ability to self-assemble into regularly structured monomolecular lattices and the functionality of the allergen

The mature crystalline bacterial cell surface (S-layer) protein SbsC of Bacillus stearothermophilus ATCC 12980 comprises amino acids 31–1099 and assembles into an oblique lattice type. As the deletion of up to 179 C-terminal amino acids did not interfere with the self-assembly properties of SbsC, th...

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Bibliographic Details
Published in:Protein engineering 2002-03, Vol.15 (3), p.243-249
Main Authors: Breitwieser, Andreas, Egelseer, Eva M., Moll, Dieter, Ilk, Nicola, Hotzy, Christoph, Bohle, Barbara, Ebner, Christof, Sleytr, Uwe B., Sára, Margit
Format: Article
Language:English
Subjects:
Allergens - genetics
Allergens - immunology
Antigens, Plant
Bacillus stearothermophilus
Bacterial Proteins - genetics
Crystallization
Geobacillus stearothermophilus
Immunoblotting
Immunoglobulin E - immunology
major birch pollen allergen
Membrane Glycoproteins - genetics
Plant Proteins - genetics
Plant Proteins - immunology
Protein Engineering
recombinant fusion protein
Recombinant Fusion Proteins - chemistry
Recombinant Fusion Proteins - genetics
Recombinant Fusion Proteins - immunology
S-layer protein
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Summary:The mature crystalline bacterial cell surface (S-layer) protein SbsC of Bacillus stearothermophilus ATCC 12980 comprises amino acids 31–1099 and assembles into an oblique lattice type. As the deletion of up to 179 C-terminal amino acids did not interfere with the self-assembly properties of SbsC, the sequence encoding the major birch pollen allergen (Bet v1) was fused to the sequence encoding the truncated form rSbsC31–920. The S-layer fusion protein, termed rSbsC/Bet v1, maintained the ability to self-assemble into flat sheets and open-ended cylinders. The presence and the functionality of the fused Bet v1 sequence was proved by blot experiments using BIP1, a monoclonal antibody against Bet v1 and Bet v1-specific IgE-containing serum samples from birch pollen allergic patients. The location and accessibility of the allergen moiety on the outer surface of the S-layer lattice were demonstrated by immunogold labeling of the rSbsC/Bet v1 monolayer, which was obtained by oriented recrystallization of the S-layer fusion protein on native cell wall sacculi. Thereby, the specific interactions between the N-terminal part of SbsC and a distinct type of secondary cell wall polymer were exploited. This is the first S-layer fusion protein described that had retained the specific properties of the S-layer protein moiety in addition to those of the fused functional peptide sequence.
ISSN:0269-2139
1741-0126
1460-213X
1741-0134
DOI:10.1093/protein/15.3.243