Abstract
BackgroundBermuda grass (Cynodon dactylon; subfamily Chloridoideae) is an important source of seasonal aeroallergens in warm tropical and sub-tropical areas worldwide. Improved approaches to diagnosis and therapy of allergic diseases require a thorough understanding of the structure and epitopes on the allergen molecule that are crucial for the antigen-antibody interaction. This study describes the localization of the human IgE-binding regions of the major group 1 pollen allergen Cyn d 1 from Bermuda grass.MethodsA cDNA library was constructed from Bermuda grass pollen (BGP) using a Lambda gt11 expression vector. The gene encoding the Cyn d 1 allergen was isolated by screening the library with a mouse monoclonal antibody raised against grass group 1 allergen. In order to characterize the IgE epitopes on Cyn d 1, seven overlapping fragments and three deletion mutants were cloned and over-expressed in E. coli. The recombinant fragments and deletion mutants were evaluated for their comparative IgE reactivity with sera of non atopic individuals and grass pollen allergic patients by ELISA and a dot-blot assay.ResultsAnalysis of IgE binding regions by overlapping fragments and deletion mutants identified two major allergenic regions corresponding to amino acids 120–170 and 224–244. Deletion of either or both regions led to a significant reduction in IgE binding, emphasizing the importance of the C-terminal region on Cyn d 1 in epitope-IgE interaction.ConclusionAnti-Cyn d 1 IgE antibodies from allergic human sera recognize two epitopes located at the C-terminal end of the molecule. These data will enable the design of improved diagnostic and therapeutic approaches for BGP hypersensitivity.
Highlights
Bermuda grass (Cynodon dactylon; subfamily Chloridoideae) is an important source of seasonal aeroallergens in warm tropical and sub-tropical areas worldwide
We report the presence of at least two major immunoglobulin E (IgE) binding regions located within the C-terminus of Cyn d 1
Results cDNA sequence analysis of Cyn d 1 A cDNA expression library was constructed from mRNA of Bermuda grass pollen (BGP) in λ gt11 vector
Summary
Bermuda grass (Cynodon dactylon; subfamily Chloridoideae) is an important source of seasonal aeroallergens in warm tropical and sub-tropical areas worldwide. This study describes the localization of the human IgE-binding regions of the major group 1 pollen allergen Cyn d 1 from Bermuda grass. Cyn d 1, the major allergen of BGP, is the most abundant protein component of BGP, comprising 15–20% of crude pollen extract. It dominates the human IgE response with 87% of individuals allergic to BGP showing positive reaction to this 32 kDa protein [6,7]. They share an N-glycosylation site at amino acid position 9 and carry glycans that account for 4–5% of their molecular weight These glycoproteins are located in the cytoplasm of the pollen grain and are rapidly released when hydrated upon contact with moist mucosal surfaces. A characteristic feature of group 1 allergens is the presence of seven strictly conserved cysteine residues, located mainly in the N-terminus of the protein [10,11,12]
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