Hypoallergenic variants of the major latex allergen Hev b 6.01 retaining human T lymphocyte reactivity.
Hev b 6.01 is a major allergen of natural rubber latex with sensitization of 70-86% of latex glove-allergic subjects. Recently, we mapped the immunodominant T cell sites of Hev b 6.01 to the highly IgE-reactive hevein (Hev b 6.02) domain. Hev b 6.01 contains 14 cysteine residues with multiple disulphide bridges stabilizing tertiary conformation. With the goal of a standardized specific immunotherapy we developed hypoallergenic Hev b 6.01 mutants by site-directed mutagenesis of selected cysteine residues (3, 12, 17, and 41) within the Hev b 6.02 domain. Peptides corresponding to the Hev b 6.02 domain of two of the mutants were also synthesized. These mutants and peptide variants showed markedly decreased or ablated latex-allergic patient serum IgE binding by immunoblotting and ELISA. Basophil activation testing confirmed markedly decreased activation with successive cysteine substitutions of the mutants and complete abrogation with the Hev b 6.02 (Cys 3, 12, 17, 41 Ala) peptide. Retention of T cell reactivity is crucial for effective specific immunotherapy and all mutants and peptide variants maintained their latex-specific T cell reactivity. The ablated allergenicity but retained T cell reactivity of the Hev b 6.02 (Cys 3, 12, 17, 41 Ala) peptide suggests this peptide is a suitable candidate for inclusion in a latex immunotherapy preparation.
Drew
Alexander C
AC
Cooperative Research Centre for Asthma, Sydney, Australia. alec.drew@med.monash.edu.au
Eusebius
Nirupama P
NP
Kenins
Linda
L
de Silva
Harini D
HD
Suphioglu
Cenk
C
Rolland
Jennifer M
JM
O'hehir
Robyn E
RE
D019943:Amino Acid Substitution / Q000235:genetics; D023181:Antimicrobial Cationic Peptides / Q000235:genetics / Q000276:immunology* / Q000378:metabolism; D001491:Basophils / Q000276:immunology / Q000378:metabolism; D001667:Binding, Competitive / Q000235:genetics / Q000276:immunology; D002455:Cell Division / Q000235:genetics / Q000276:immunology; D002460:Cell Line; D003888:Desensitization, Immunologic / Q000379:methods; D004220:Disulfides / Q000737:chemistry; D015536:Down-Regulation; D018984:Epitopes, T-Lymphocyte / Q000235:genetics / Q000276:immunology / Q000378:metabolism; D006801:Humans; D007073:Immunoglobulin E / Q000096:biosynthesis / Q000378:metabolism; D020315:Latex Hypersensitivity / Q000175:diagnosis / Q000276:immunology*; D008213:Lymphocyte Activation / Q000235:genetics / Q000276:immunology; D010446:Peptide Fragments / Q000235:genetics / Q000276:immunology / Q000378:metabolism; D037121:Plant Lectins / Q000235:genetics / Q000276:immunology* / Q000378:metabolism; D011485:Protein Binding / Q000235:genetics / Q000276:immunology; D017434:Protein Structure, Tertiary / Q000235:genetics; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000235:genetics / Q000276:immunology; D013601:T-Lymphocytes / Q000166:cytology / Q000276:immunology*
D023181:Antimicrobial Cationic Peptides; D004220:Disulfides; D018984:Epitopes, T-Lymphocyte; D010446:Peptide Fragments; D037121:Plant Lectins; D011994:Recombinant Proteins; C045875:hevein; D007073:Immunoglobulin E