Involvement of the N-terminus of Der p 2 in IgE and monoclonal antibody binding.
The major house dust mite allergen Der p 2 was expressed as a recombinant fusion protein in Escherichia coli either with glutathione-S-transferase as fusion partner or with a poly-histidine tag. Both recombinant fusion proteins failed to react with 3/14 Der p 2-specific monoclonal antibodies (mAbs). When Der p 2 was expressed in yeast with one alanine linked N-terminally to the allergen, no reactivity was observed. When expressed without any fusion partner, all 14 mAbs showed reactivity. The addition of a single N-terminal alanine also disrupted an important epitope for IgE. In RAST inhibitions, an average decrease in inhibitory potency of 72+/-32% was observed (n = 16) with a maximum decrease of 91%. These observations suggest that the N-terminus of Der p 2 is involved in an important epitope for IgE that is disrupted by the addition of one single amino-acid. Recombinant Der p 2 molecules should therefore preferably lack any fusion peptide.
Hakkaart
G A
GA
Central Laboratory of the Netherlands Red Cross Blood Transfusion Service, University of Amsterdam, The Netherlands.
Aalberse
R C
RC
van Ree
R
R
D000409:Alanine; D000485:Allergens / Q000235:genetics / Q000276:immunology*; D000818:Animals; D000911:Antibodies, Monoclonal / Q000276:immunology*; D000941:Antigens / Q000235:genetics / Q000276:immunology; D039741:Antigens, Dermatophagoides; D001666:Binding Sites, Antibody / Q000276:immunology*; D004797:Enzyme-Linked Immunosorbent Assay; D000939:Epitopes / Q000276:immunology; D004926:Escherichia coli / Q000235:genetics; D015870:Gene Expression; D006023:Glycoproteins / Q000235:genetics / Q000276:immunology*; D006801:Humans; D007073:Immunoglobulin E / Q000276:immunology*; D051379:Mice; D008807:Mice, Inbred BALB C; D008925:Mites / Q000276:immunology*; D010957:Plasmids; D011852:Radioallergosorbent Test; D011993:Recombinant Fusion Proteins / Q000276:immunology; D012441:Saccharomyces cerevisiae / Q000235:genetics
D000485:Allergens; D000911:Antibodies, Monoclonal; D000941:Antigens; D039741:Antigens, Dermatophagoides; D000939:Epitopes; D006023:Glycoproteins; D011993:Recombinant Fusion Proteins; D007073:Immunoglobulin E; D000409:Alanine