title: Affinity improvement of the high-affinity immunoglobulin E receptor by phage display.
abstract: The immunoglobulin E (IgE)-binding site of its high-affinity receptor is localized in the second immunoglobulin-like domain (D2) of the alpha-subunit (Fc epsilon RI alpha). In this study, the randomized pentapeptides were introduced between Glu(132) and Ile(138) of Fc epsilon RI alpha D2 and displayed on a filamentous phage. After eight rounds of panning, a phage clone having a mutation of Asp(135)Tyr(136)Met(137) in Fc epsilon RI alpha D2 was obtained. The binding affinity of the mutant phages to immobilized IgE was approximately 500 times higher than that of the wild type. The mutant phages competitively inhibited the binding of IgE to the soluble receptor at a 50% inhibition (IC(50)) value of 116 pM. The mutant Fc epsilon RI alpha D2, which had been expressed as a fusion protein with glutathione S-transferase in Escherichia coli, also showed higher IgE-binding capacity than the wild type. The mutant Fc epsilon RI alpha D2 is expected to manifest its improved IgE-binding affinity together with any fusion partner.
authors.affiliation: Institute of Applied Biochemistry, University of Tsukuba, 1-1-1 Tennodai, Tsukuba, Ibaraki 305-8572, Japan.
authors.forename: Akio
authors.initials: A
authors.lastname: Iwasaki
authors.forename: Takeshi
authors.initials: T
authors.lastname: Doi
authors.forename: Michihisa
authors.initials: M
authors.lastname: Umetani
authors.forename: Masanao
authors.initials: M
authors.lastname: Watanabe
authors.forename: Makoto
authors.initials: M
authors.lastname: Suda
authors.forename: Yukio
authors.initials: Y
authors.lastname: Hattori
authors.forename: Takao
authors.initials: T
authors.lastname: Nagoya
mesh_terms: D000595:Amino Acid Sequence; D019943:Amino Acid Substitution; D001483:Base Sequence; D001665:Binding Sites; D003001:Cloning, Molecular; D004926:Escherichia coli / Q000235:genetics; D006801:Humans; D007073:Immunoglobulin E / Q000378:metabolism*; D007700:Kinetics; D008969:Molecular Sequence Data; D016296:Mutagenesis; D019151:Peptide Library; D016133:Polymerase Chain Reaction; D017455:Receptors, IgE / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism*; D011993:Recombinant Fusion Proteins / Q000378:metabolism
chemical_list: D019151:Peptide Library; D017455:Receptors, IgE; D011993:Recombinant Fusion Proteins; D007073:Immunoglobulin E