Using Monoclonal Antibodies to Characterize a Sequential Epitope on the Group I Allergen of Bermuda Grass Pollen

Abstract

Background: Cyn d 1, the group I allergen of Bermuda grass pollen, had been purified and characterized. Methods: A sequential B cell epitope on Cyn d 1 was studied with monoclonal antibodies (MoAbs). Cyn d 1 was cleaved by Achromobacter protease I into fragments, and the resulting peptides were fractionated on reversed-phase columns before being reacted with anti-Cyn d 1 MoAbs in a radioimmunoassay. A Cyn d 1 fragment recognized by its MoAb was selected for Edman degradation. A synthetic peptide was constructed according to the determined sequence. Results: The epitope on Cyn d 1 recognized by MoAb 18–53 was found to be conformation independent, since its activity was not changed after sodium periodate, guanidine or urea treatment. The enzyme-cleaved fragment containing this epitope was determined to be DVDKPPFDGMTACGNEPIF which corresponds to the N-terminal 46–64 residues of Cyn d 1. The presence of this sequence in the epitope recognized by MoAb 18–53 was demonstrated by enzyme immunoassay and further confirmed by inhibition of binding enzyme immunoassay with synthetic peptides. Some cross-reactivity with the N-terminal 45–63 residues of Lol p 1 was also found. Conclusions: The primary structure of a sequential epitope on Cyn d 1 was determined, and its activity was confirmed with peptides synthesized according to the determined sequence.

Sequence polymorphism of the group 1 allergen of Bermuda grass pollen

“…MoAb 4-37 is against an epitope common to all isoforms of Cyn d 1 [5], and MoAb 18-53 has recently been found to be against a sequential epitope on the 46-64th residues of Cyn d 1 [10]. The cDNA library was constructed from BGP obtained via two different resources, and the cDNA clones were screened by immunoassay with a mixture of MoAbs 4-37 and 18-53.…”

“…The cDNA library was constructed from BGP obtained via two different resources, and the cDNA clones were screened by immunoassay with a mixture of MoAbs 4-37 and 18-53. MoAb 4-37 is against an epitope common to all isoforms of Cyn d 1 [5], and MoAb 18-53 has recently been found to be against a sequential epitope on the 46-64th residues of Cyn d 1 [10]. All the 10 cDNA clones encode the full length of Cyn d 1 (Fig.…”

Induction of Specific Th1 Responses and Suppression of IgE Antibody Formation by Vaccination with Plasmid DNA Encoding Cyn d 1

“…The sequences of some cDNA clones of Cyn d 1 were first identified by Smith et al [6]. However, the cDNAs were truncated in the coding regions [7]. Several full-length cDNA clones encoding Cyn d 1 were thereafter characterized by us [8,9].…”

“…Most patients allergic to BGP have IgE antibodies specific for Cyn d 1 [5,6,7,8]. Smith et al [6] partially characterized cDNA clones encoding Cyn d 1, but the N-terminal amino acid sequence of Cyn d 1 was cloned by us [7]. Recombinant Cyn d 1 (rCyn d 1) was used to set up a mouse allergic model in this study.…”

Mapping of IgE and IgG₄ Antibody-Binding Epitopes in Cyn d 1, the Major Allergen of Bermuda Grass Pollen

“…Using mouse monoclonal or rabbit polyclonal antibodies, a number of antibody-binding epitopes have been disclosed for aeroallergens and food allergens [17,18,19,20,21,22,23]. We have previously characterized a sequential epitope (peptides 46–64) on Cyn d 1, the major allergen of BGP [24]. Synthetic peptides have been recently applied for epitope mapping [25,26,27,28].…”