Abstract
BackgroundThe interaction between IgE and allergen is a key event at the initiation of an allergic response, and its characteristics have substantial effects on the clinical manifestation. Despite this, the molecular details of the interaction between human IgE and the major birch allergen Bet v 1, one of the most potent tree allergens, still remain poorly investigated.ObjectiveTo isolate Bet v 1-specific human monoclonal IgE and characterize their interaction with the allergen.MethodsRecombinant human IgE were isolated from a combinatorial antibody fragment library and their interaction with Bet v 1 assessed using various immunological assays. The structure of one such IgE in the single-chain fragment variable format was determined using X-ray crystallography.ResultsWe present four novel Bet v 1-specific IgE, for one of which we solve the structure, all with their genetic origin in the IGHV5 germline gene, and demonstrate that they target two non-overlapping epitopes on the surface of Bet v 1, thereby fulfilling the basic criteria for FcεRI cross-linkage. We further define these epitopes and for one epitope pinpoint single amino acid residues important for the interaction with human IgE. This provides a potential explanation, at the molecular level, for the differences in recognition of isoforms of Bet v 1 and other allergens in the PR-10 protein family displayed by IgE targeting this epitope. Finally, we present the first high-resolution structure of a human allergen-specific IgE fragment in the single-chain fragment variable (scFv) format.Conclusions and Clinical RelevanceWe here display the usefulness of allergen-specific human monoclonal IgE as a tool in studies of the crucial molecular interaction taking place at the initiation of an allergic response. Such studies may aid us in development of better diagnostic tools and guide us in the development of new therapeutic compounds.
Highlights
A type I hypersensitivity response is initiated when allergen-specific IgE bound to high-affinity IgE receptors (FceRI) on the surface of effector cells, such as mast cells and basophils, are cross-linked by allergen
An overrepresentation of IgE-encoding transcripts derived from the immunoglobulin heavy variable (IGHV) 5 germline gene subgroup has been observed in such tissue samples [4,5,6]
We here approach these issues and present a molecular characterization of Bet v 1-specific human antibody fragments derived from the IgE repertoire of allergic individuals. These antibody fragments have a genetic origin in IGHV5, the germline gene subgroup sometimes implicated as overrepresented in IgE repertoires [4,5,6]. We show that such a repertoire of clonally different antibodies, despite their origin in a common immunoglobulin VH gene, is able to target more than one epitope on Bet v 1, thereby in principle fulfilling the prerequisites for crosslinkage and initiation of the allergic cascade
Summary
A type I hypersensitivity response is initiated when allergen-specific IgE bound to high-affinity IgE receptors (FceRI) on the surface of effector cells, such as mast cells and basophils, are cross-linked by allergen. This triggers a cascade of events eventually giving rise to the release of biologically active compounds that exert their effects both locally and systemically [1, 2]. One intriguing outcome of several of the past IgE-repertoire studies is a significant difference in the utilization of different immunoglobulin germline gene subgroups to produce the heavy chain variable (VH) domains of IgE in certain tissue as compared to IgE and other isotypes encoded by peripheral blood lymphocytes. Two possible explanations are that such skewed repertoires are the result of either a polyclonal expansion of B cells expressing surface IgE with origin in these genes, for example, by bacterial superantigens [14] or of a clonal selection process where certain allergens favour the selection of clones originating in a limited set of germline genes [10]
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