Interaction of human IgE with Fc epsilon RI alpha exposes hidden epitopes on IgE.
BACKGROUND
Binding of human IgE via the heavy-chain constant region domain 3 (Cepsilon3) to the alpha-chain of its high affinity receptor (FcepsilonRIalpha) is a key event in mediating allergic reactions. We wanted to identify epitopes within Cepsilon3 that are stable to denaturation and to evaluate whether such structures are involved in receptor binding. The existence of stable epitopes would facilitate the generation of compounds that inhibit the IgE-FcepsilonRIalpha interaction.


METHODS
Monoclonal anti-human IgE-antibodies against recombinant bacterially synthesized Cepsilon3, which is known to be partly misfolded, were raised in mice. These antibodies were probed for binding to native, immobilized and receptor-bound IgE, respectively, providing tools for the identification of the indicated stable epitopes.


RESULTS
Two of the generated antibodies (8E7, 3G9) discriminate between IgE in solution and IgE attached to FcepsilonRIalpha, pointing towards a steric rearrangement within Cepsilon3 induced upon receptor binding. The described antibodies represent tools for studying the mechanism of the Fcepsilon-FcepsilonRIalpha interaction and may be of diagnostic value since serum IgE from various human donors was differently recognized by 8E7, which is indicative for naturally occurring IgE molecules with different steric conformation.


CONCLUSIONS
The presented data support the hypothesis of a conformational change within IgE Cepsilon3 upon receptor binding by showing that monoclonal antibodies raised against recombinant Cepsilon3 differently recognize soluble and receptor-bound IgE. The presence of an IgE portion in sera of human donors that is recognized by 8E7 indicates the existence of IgE molecules in different steric conformations in human blood, which may be related to pathologic parameters.
Nechansky
A
A
Novartis Forschungsinstitut GmbH, Vienna, Austria.
Ruf
C
C
Pursch
E
E
Wasserbauer
E
E
Stämpfli
M R
MR
Pachlopnik
J M
JM
Stadler
B M
BM
Kricek
F
F
D000911:Antibodies, Monoclonal / Q000276:immunology; D000917:Antibody Formation; D000939:Epitopes / Q000276:immunology; D006801:Humans; D007073:Immunoglobulin E / Q000276:immunology* / Q000378:metabolism; D007141:Immunoglobulin Fc Fragments / Q000276:immunology; D011489:Protein Denaturation; D017455:Receptors, IgE / Q000378:metabolism*; D011994:Recombinant Proteins / Q000276:immunology
D000911:Antibodies, Monoclonal; D000939:Epitopes; D007141:Immunoglobulin Fc Fragments; D017455:Receptors, IgE; D011994:Recombinant Proteins; D007073:Immunoglobulin E