title: Construction of affinity changeable antibody in response to Ca2+.
abstract: Immunoaffinity chromatography is a powerful method for purification of proteins because of the high selectivity and avidity of antibodies. Due to the strength of antigen-antibody binding, however, elution of proteins bound to antibodies that are covalently immobilized on the column is performed by temporary denaturation of the antibody. Therefore, the development of milder elution conditions could improve the recovery of the antibodies and prolong the life of the immunoaffinity column. We describe the design and construction of an antibody that changes its affinity in response to external stimuli. The heavy chain and light chain of a single chain Fv of the D1.3 antibody against hen egg-white lysozyme (HEL) were fused at the N- and C-termini, respectively, of the calmodulin-M13 fusion protein. The affinity of this fusion protein for HEL could be modulated by changing the Ca(2+) concentration.
authors.affiliation: Department of Biological Information, Graduate School of Bioscience and Biotechnology, Tokyo Institute of Technology, Midori-ku, Yokohama, Japan. ekobatak@bio.titech.ac.jp
authors.forename: Eiry
authors.initials: E
authors.lastname: Kobatake
authors.forename: Chihiro
authors.initials: C
authors.lastname: Kosaku
authors.forename: Satoshi
authors.initials: S
authors.lastname: Hanzawa
authors.forename: Masayasu
authors.initials: M
authors.lastname: Mie
mesh_terms: D000915:Antibody Affinity / Q000187:drug effects*; D002118:Calcium / Q000378:metabolism*; D002413:Cations, Divalent / Q000378:metabolism*; D002846:Chromatography, Affinity / Q000379:methods; D009113:Muramidase / Q000302:isolation & purification; D011994:Recombinant Proteins / Q000235:genetics / Q000276:immunology / Q000378:metabolism; D057127:Single-Chain Antibodies / Q000235:genetics / Q000276:immunology* / Q000378:metabolism*
chemical_list: D002413:Cations, Divalent; D011994:Recombinant Proteins; D057127:Single-Chain Antibodies; C421864:hen egg lysozyme; D009113:Muramidase; D002118:Calcium