title: The molecular basis of antigenic cross-reactivity between the group 2 mite allergens.
abstract: BACKGROUND
  Mite group 2 allergens Der p 2, Der f 2, and Eur m 2 are 14-kDa proteins of unknown function that share 83% to 85% amino acid sequence identity. Isoforms of the allergens within each genus have been identified which differ by 3 or 4 amino acids, but little is known of the influence of group 2 polymorphisms on human IgE antibody binding.
  
  
  OBJECTIVE
  The purpose of this study was to investigate the importance of interspecies and isoform substitutions on murine mAb and IgE antibody binding and on the molecular structure of the group 2 allergens.
  
  
  METHODS
  Site-directed mutagenesis was used to incorporate the isoform amino acid substitutions onto the Der p 2.0101 sequence. Recombinant allergens were expressed and purified from Escherichia coli and used to evaluate antibody binding by enzyme-linked immunosorbent assay (ELISA). Molecular modeling of the tertiary structure was used to analyze structural differences between the various group 2 allergens.
  
  
  RESULTS
  The substitution of asparagine for aspartic acid at position 114 restored mAb binding of rDer p 2.0101; the other Der p 2 isoforms and the 3 rDer f 2 isoforms also reacted in the 2-site ELISA. The correlation of IgE binding to the Der p 2 isoforms was excellent and tended to be higher in the isoforms with the asparagine 114 substitution (r (2) = 0.87 vs r (2) = 0.95). rEur m 2.0101 bound to all mAb except 7A1; when compared with rDer p 2 for IgE binding, rEur m 2.0101 gave a correlation coefficient of r (2) = 0.68. Molecular modeling revealed that Eur m 2 and the storage mite homologs Lep d 2 and Tyr p 2 retain the tertiary fold of Der p 2. Eur m 2 has a conserved surface, whereas Lep d 2 and Tyr p 2 present most of the amino acid substitutions on this surface. Lep d 2 and Tyr p 2 did not react with mAb or with sera from patients with IgE to Dermatophagoides species.
  
  
  CONCLUSIONS
  The isoform substitutions of rDer p 2 can be distinguished by mAb. The allergenic cross-reactivity between Der p 2, Der f 2, and Eur m 2 is a direct result of the conserved antigenic surface, whereas the lack of cross-reactivity with Lep d 2 and Tyr p 2 is a result of the multiple substitutions across this surface.
authors.affiliation: Asthma & Allergic Diseases Center, Department of Medicine, University of Virginia Health System, Charlottesville, VA 22908-1355, USA.
authors.forename: A M
authors.initials: AM
authors.lastname: Smith
authors.forename: D C
authors.initials: DC
authors.lastname: Benjamin
authors.forename: N
authors.initials: N
authors.lastname: Hozic
authors.forename: U
authors.initials: U
authors.lastname: Derewenda
authors.forename: W A
authors.initials: WA
authors.lastname: Smith
authors.forename: W R
authors.initials: WR
authors.lastname: Thomas
authors.forename: G
authors.initials: G
authors.lastname: Gafvelin
authors.forename: M
authors.initials: M
authors.lastname: van Hage-Hamsten
authors.forename: M D
authors.initials: MD
authors.lastname: Chapman
mesh_terms: D000595:Amino Acid Sequence; D019943:Amino Acid Substitution; D000818:Animals; D000911:Antibodies, Monoclonal / Q000276:immunology* / Q000378:metabolism; D039741:Antigens, Dermatophagoides; D003429:Cross Reactions; D000939:Epitopes / Q000276:immunology*; D006023:Glycoproteins / Q000737:chemistry / Q000235:genetics* / Q000276:immunology* / Q000378:metabolism; D006801:Humans; D006969:Hypersensitivity, Immediate / Q000276:immunology; D007073:Immunoglobulin E / Q000378:metabolism; D008925:Mites / Q000276:immunology*; D008958:Models, Molecular; D008969:Molecular Sequence Data; D020033:Protein Isoforms / Q000737:chemistry / Q000378:metabolism; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism
chemical_list: D000911:Antibodies, Monoclonal; D039741:Antigens, Dermatophagoides; D000939:Epitopes; D006023:Glycoproteins; D020033:Protein Isoforms; D011994:Recombinant Proteins; D007073:Immunoglobulin E