Preserved epitope-specific T cell activation by recombinant Bet v 1-MBP fusion proteins.
BACKGROUND
Allergen-specific T cells play an important role in the allergic immune response, and are thought to be the principal target in specific immunotherapy.


OBJECTIVE
The aim of the present study was to evaluate if fusion proteins of allergens with bacterial proteins can be used to activate and bias allergen-specific T cells, and to characterize T cell epitopes.


METHODS
The complete gene of Bet v 1, the major birch pollen allergen, was amplified by PCR from birch pollen mRNA, and cloned in pKK223-3. The complete gene or truncated sequences were transferred to pMAL-c and expressed in E. coli as fusion proteins with maltose binding protein (MBP). The complete fusion protein, and the truncated fusion proteins were used for studies with Bet v 1-specific T cells.


RESULTS
Bet v 1-specific T cells reacted similarly with purified and crude Bet v 1-MBP proteins. Therefore, crude preparations were used to study the epitope-specificity of 11 Bet v 1-specific T cell clones. Six distinct T cell epitopes were determined in this way. Interestingly, the T cell epitope of three T cell clones, that did not react with synthetic peptides in a previous study, was identified. In addition, the presence of MBP as a fusion partner to Bet v 1 was shown to influence TH2/TH1 cytokine production in T cell lines, but not in established T cell clones.


CONCLUSIONS
Using crude preparations of recombinant fusion proteins of Bet v 1 with MBP, multiple T cell epitopes were identified in Bet v 1. As T cell activation is preserved in this system, the generation of recombinant allergens with TH1-inducing proteins as fusion partners might be considered as a T-cell targeted approach for specific immunotherapy.
van Neerven
R J
RJ
ALK ABELLO Group, Hørsholm, Denmark.
Sparholt
S H
SH
Schou
C
C
Larsen
J N
JN
D018528:ATP-Binding Cassette Transporters; D000485:Allergens; D000595:Amino Acid Sequence; D052179:Antigens, Plant; D001426:Bacterial Proteins / Q000235:genetics / Q000276:immunology; D002352:Carrier Proteins / Q000235:genetics / Q000276:immunology*; D002460:Cell Line / Q000737:chemistry / Q000276:immunology / Q000378:metabolism; D002999:Clone Cells / Q000737:chemistry / Q000276:immunology / Q000378:metabolism; D003001:Cloning, Molecular; D016207:Cytokines / Q000276:immunology / Q000378:metabolism; D000939:Epitopes / Q000737:chemistry / Q000235:genetics / Q000276:immunology*; D029968:Escherichia coli Proteins; D005798:Genes, Bacterial / Q000235:genetics; D017343:Genes, Plant / Q000235:genetics; D006801:Humans; D008213:Lymphocyte Activation / Q000276:immunology*; D058619:Maltose-Binding Proteins; D008969:Molecular Sequence Data; D009004:Monosaccharide Transport Proteins; D010940:Plant Proteins / Q000737:chemistry / Q000235:genetics / Q000276:immunology*; D010957:Plasmids / Q000235:genetics; D011993:Recombinant Fusion Proteins / Q000737:chemistry / Q000235:genetics / Q000276:immunology; D013601:T-Lymphocytes / Q000166:cytology / Q000276:immunology* / Q000378:metabolism
D018528:ATP-Binding Cassette Transporters; D000485:Allergens; D052179:Antigens, Plant; D001426:Bacterial Proteins; D002352:Carrier Proteins; D016207:Cytokines; D000939:Epitopes; D029968:Escherichia coli Proteins; D058619:Maltose-Binding Proteins; D009004:Monosaccharide Transport Proteins; D010940:Plant Proteins; D011993:Recombinant Fusion Proteins; C461599:maltose transport system, E coli; C061312:Bet v 1 allergen, Betula