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Characterization of B‐ and T‐cell responses and HLA‐DR4 binding motifs of the latex allergen Hev b 6.01 (prohevein) and its post‐transcriptionally formed proteins Hev b 6.02 and Hev b 6.03
Author(s) -
RaulfHeimsoth M.,
Rozynek P.,
Brüning T.,
Rihs H.P.
Publication year - 2004
Publication title -
allergy
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 3.363
H-Index - 173
eISSN - 1398-9995
pISSN - 0105-4538
DOI - 10.1111/j.1398-9995.2004.00475.x
Subject(s) - immunoglobulin e , maltose binding protein , latex allergy , immunology , recombinant dna , peripheral blood mononuclear cell , antibody , b cell , biology , chemistry , allergy , fusion protein , in vitro , biochemistry , gene
Background: Multiple immunoglobulin E (IgE)‐binding proteins in natural rubber latex extracts have been identified. In the case of Hev b 6 a differentiation was made between the precursor protein prohevein (Hev b 6.01) and its two post‐transcriptionally formed proteins, the N‐terminal hevein (Hev b 6.02) and the C‐terminal domain (Hev b 6.03). All three components act as independent allergens. The aim of this study was a detailed analysis of the T‐cell responses and the IgE‐binding capacity of Hev b 6.01, Hev b 6.02 and Hev b 6.03 by using these allergens as recombinant maltose‐binding fusion (MBP) proteins and the usage of synthetic modified hevein peptides. Methods: Latex‐allergic health care workers (HCWs) suffering from rhinitis, conjunctivitis, contact urticaria and/or asthma with increased specific IgE‐antibodies to latex were tested for their IgE‐binding capacity and T‐cell reactivity (by proliferation response) to the recombinant MBP‐rHev b 6.01, MBP‐rHev b 6.02, MBP‐rHev b 6.03, to native Hev b 6.02, to modified hevein peptides and wheat germ agglutinin (WGA). For testing of the human leucocyte antigen (HLA) class II restriction of MBP‐rHev b 6.01 induced peripheral blood mononuclear cell (PBMC) responses, monoclonal antibodies against HLA‐DR, HLA‐DP or HLA‐DQ were added. Results: Seventeen of 18 (94%) serum samples from latex‐allergic HCWs had increased levels of specific IgE to MBP‐rHev b 6.01, 16 (89%) to MBP‐rHev b 6.02 and 13 (72%) to MBP‐rHev b 6.03. A significant difference existed between the specific IgE‐values of MBP‐rHev b 6.02 and MBP‐rHev b 6.03 ( P < 0.01). Proliferation responses of PBMC of the same 18 latex‐allergic patients were positive for MBP‐rHev b 6.01 and MBP‐rHev b 6.03 in 83 and 67% of the tested PBMC suspension, whereas the proliferation responses induced with MBP‐rHev b 6.02 or native Hev b 6.02 were very low (5.6 and 22.2%). Sera from nine additional latex‐allergic patients showed specific IgE binding to the native Hev b 6.02, but none of these sera showed specific IgE binding to the modified Hev b 6.02‐peptides [whereby all eight cysteine residues were substituted by serine (C → S) or by alanine (C → A)]. Proliferation responses induced by the modified Hev b 6.02 peptides were not significantly different from that induced by Hev b 6.02. Potential HLA‐DR4Dw4(DRB1*0401)‐restricted T‐cell epitopes of Hev b 6.01 predicted by two computer algorithms were only found in the Hev b 6.03‐part of Hev b 6.01. Conclusion: In the Hev b 6.01 precursor the regions responsible for IgE binding and those for inducing the T‐cell proliferation responses are settled in different parts of the protein. The Hev b 6.02 domain is responsible for IgE binding and carries discontinuous B‐cell epitopes whereas Hev b 6.03 is a better inducer of a proliferation response and contains HLA‐DR4‐binding motifs.