Mutagenesis within human FcepsilonRIalpha differentially affects human and murine IgE binding.
Soluble fragments of the alpha-chain of FcepsilonRI, the high-affinity receptor for IgE, compete with membrane-bound receptors for IgE and may thus provide a means to combat allergic responses. Mutagenesis within FcepsilonRIalpha is used in this study, in conjunction with the crystal structure of the FcepsilonRIalpha/IgE complex, to define the relative importance of specific residues within human FcepsilonRIalpha for IgE binding. We have also compared the effects of these mutants on binding to both human and mouse IgE, with a view to evaluating the mouse as an appropriate model for the analysis of future agents designed to mimic the human FcepsilonRIalpha and attenuate allergic disease. Three residues within the C-C' region of the FcepsilonRIalpha2 domain and two residues within the alpha2 proximal loops of the alpha1 domain were selected for mutagenesis and tested in binding assays with human and mouse IgE. All three alpha2 mutations (K117D, W130A, and Y131A) reduced the affinity of human IgE binding to different extents, but K117D had a far more pronounced effect on mouse IgE binding, and although Y131A had little effect, W130A modestly enhanced binding to mouse IgE. The mutations in alpha1 (R15A and F17A) diminished binding to both human and mouse IgE, with these effects most likely caused by disruption of the alpha1/alpha2 interface. Our results demonstrate that the effects of mutations in human FcepsilonRIalpha on mouse IgE binding, and hence the inhibitory properties of human receptor-based peptides assayed in rodent models of allergy, may not necessarily reflect their activity in a human IgE-based system.
Mackay
Graham A
GA
Randall Centre, New Hunt's House, King's College London, Guy's Campus, London, United Kingdom.
Hulett
Mark D
MD
Cook
Justin P D
JP
Trist
Halina M
HM
Henry
Alistair J
AJ
McDonnell
James M
JM
Beavil
Andrew J
AJ
Beavil
Rebecca L
RL
Sutton
Brian J
BJ
Hogarth
P Mark
PM
Gould
Hannah J
HJ
D000818:Animals; D001491:Basophils / Q000276:immunology; D015550:Cell Degranulation; D002942:Circular Dichroism; D006636:Histamine Release; D006801:Humans; D007073:Immunoglobulin E / Q000276:immunology / Q000378:metabolism*; D007700:Kinetics; D051379:Mice; D008958:Models, Molecular; D016297:Mutagenesis, Site-Directed; D010843:Pichia / Q000235:genetics; D017434:Protein Structure, Tertiary; D051381:Rats; D017455:Receptors, IgE / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism*; D011994:Recombinant Proteins / Q000378:metabolism; D013045:Species Specificity; D020349:Surface Plasmon Resonance; D014407:Tumor Cells, Cultured
D017455:Receptors, IgE; D011994:Recombinant Proteins; D007073:Immunoglobulin E