Human Fc epsilon RIalpha-specific human single-chain Fv (scFv) antibody with antagonistic activity toward IgE/Fc epsilon RIalpha-binding.
The alpha-chain of Fc epsilon RI (Fc epsilon RIalpha) plays a critical role in the binding of IgE to Fc epsilon RI. A fully human antibody interfering with this interaction may be useful for the prevention of IgE-mediated allergic diseases. Here, we describe the successful isolation of a human single-chain Fv antibody specific to human Fc epsilon RIalpha using human antibody phage display libraries. Using the non-immune phage antibody libraries constructed from peripheral blood lymphocyte cDNA from 20 healthy subjects, we isolated three phage clones (designated as FcR epsilon 27, FcR epsilon 51, and FcR epsilon 70) through two rounds of biopanning selection. The purified soluble scFv, FcR epsilon 51, inhibited the binding of IgE to recombinant Fc epsilon RIalpha, although both FcR epsilon 27 and FcR epsilon 70 showed fine binding specificity to Fc epsilon RIalpha. Since FcR epsilon 51 was determined to be a monomer by HPLC, BIAcore analysis was performed. The dissociation constant of FcR epsilon 51 to Fc epsilon RIalpha was estimated to be 20 nM, i.e., fortyfold lower than that of IgE binding to Fc epsilon RIalpha (K(d) = 0.5 nM). With these characteristics, FcR epsilon 51 exhibited inhibitory activity on the release of histamine from passively sensitized human peripheral blood mononuclear cells.
Hashiguchi
Shuhei
S
Department of Bioengineering, Faculty of Engineering, Kagoshima University, 1-21-40 Korimoto, Kagoshima 890-0065.
Nakashima
Toshihiro
T
Nitani
Aya
A
Yoshihara
Tomoki
T
Yoshinaga
Keisuke
K
Ito
Yuji
Y
Maeda
Yoshio
Y
Sugimura
Kazuhisa
K
D000595:Amino Acid Sequence; D015153:Blotting, Western; D002478:Cells, Cultured; D006636:Histamine Release / Q000187:drug effects; D006801:Humans; D006967:Hypersensitivity / Q000628:therapy; D007073:Immunoglobulin E / Q000276:immunology / Q000378:metabolism*; D007141:Immunoglobulin Fc Fragments / Q000737:chemistry; D007135:Immunoglobulin Variable Region / Q000737:chemistry / Q000235:genetics / Q000276:immunology / Q000494:pharmacology*; D007963:Leukocytes, Mononuclear / Q000276:immunology; D008969:Molecular Sequence Data; D019151:Peptide Library; D017455:Receptors, IgE / Q000037:antagonists & inhibitors* / Q000276:immunology / Q000378:metabolism; D016415:Sequence Alignment; D057127:Single-Chain Antibodies
C478757:FcRepsilon51 protein, human; D007141:Immunoglobulin Fc Fragments; D007135:Immunoglobulin Variable Region; D019151:Peptide Library; D017455:Receptors, IgE; D057127:Single-Chain Antibodies; D007073:Immunoglobulin E