Identification of sites on the human Fc epsilon RI alpha subunit which are involved in binding human and rat IgE.
The high affinity IgE Fc receptor (Fc epsilon RI), found on mast cells and basophils, is a tetrameric receptor complex. The extracellular portion of the Fc epsilon RI alpha subunit consists of two immunoglobulin-like domains and binds IgE in the absence of the other subunits. To localize the high affinity IgE binding site within the Fc epsilon RI alpha subunit, we generated a series of chimeric receptor constructs where one of the two immunoglobulin-like domains was either deleted or substituted with those from the human Fc gamma RIIIA alpha or the rat Fc epsilon RI alpha subunit. The chimeric receptors were monitored for their capacity to bind human and rat IgE, and their reactivity with different antireceptor antibodies. Domain I substitutions maintained high affinity human IgE binding. Domain II substitutions resulted in a total loss of both human and rat IgE binding. Single-domain alpha subunits could not bind IgE, suggesting that both extracellular domains are required for proper protein folding or IgE binding. To further localize the IgE binding sites, homolog-scanning mutagenesis was performed. At least three independent regions of domain II encompassing residues 118-129, 136-150, and 148-162 were required for IgE binding. Our results suggest that domain II of the human Fc epsilon RI alpha confers most of the important contributions to the binding of the human IgE Fc molecule, whereas domain I of the rat Fc epsilon RI alpha makes important contributions to the binding of rat IgE.
Mallamaci
M A
MA
Department of Bronchopulmonary Research, Hoffmann-La Roche Inc., Nutley, New Jersey 07110.
Chizzonite
R
R
Griffin
M
M
Nettleton
M
M
Hakimi
J
J
Tsien
W H
WH
Kochan
J P
JP
D000595:Amino Acid Sequence; D000818:Animals; D001665:Binding Sites; D002478:Cells, Cultured; D006801:Humans; D007073:Immunoglobulin E / Q000378:metabolism*; D008969:Molecular Sequence Data; D016296:Mutagenesis; D051381:Rats; D017455:Receptors, IgE / Q000737:chemistry / Q000378:metabolism*; D011993:Recombinant Fusion Proteins / Q000378:metabolism
D017455:Receptors, IgE; D011993:Recombinant Fusion Proteins; D007073:Immunoglobulin E