title: Identification of small molecule binding molecules by affinity purification using a specific ligand immobilized on PEGA resin.
abstract: We investigated the application of resins used in solid-phase synthesis for affinity purification. A synthetic ligand for FK506-binding protein 12 (SLF) was immobilized on various resins, and the binding assays between the SLF-immobilized resins and FK506-binding protein 12 (FKBP12) were performed. Of the resins tested in this study, PEGA resin was the most effective for isolating FKBP12. This matrix enabled the isolation of FKBP12 from a cell lysate, and the identification of SLF-binding peptides from a phage cDNA library. We confirmed the interaction between SLF and these peptides using a cuvette type quartz crystal microbalance (QCM) apparatus. Our study suggests that PEGA resin has great potential as a tool not only for the purification and identification of small-molecule binding proteins but also for the selection of peptides that recognize target molecules.
authors.affiliation: Department of Applied Biological Science, Tokyo University of Science, 2641 Yamazaki, Noda, Chiba 278-8510, Japan.
authors.forename: Kouji
authors.initials: K
authors.lastname: Kuramochi
authors.forename: Yuka
authors.initials: Y
authors.lastname: Miyano
authors.forename: Yoshihiro
authors.initials: Y
authors.lastname: Enomoto
authors.forename: Ryo
authors.initials: R
authors.lastname: Takeuchi
authors.forename: Kazutomo
authors.initials: K
authors.lastname: Ishi
authors.forename: Yoichi
authors.initials: Y
authors.lastname: Takakusagi
authors.forename: Takeki
authors.initials: T
authors.lastname: Saitoh
authors.forename: Keishi
authors.initials: K
authors.lastname: Fukudome
authors.forename: Daisuke
authors.initials: D
authors.lastname: Manita
authors.forename: Yoshifumi
authors.initials: Y
authors.lastname: Takeda
authors.forename: Susumu
authors.initials: S
authors.lastname: Kobayashi
authors.forename: Kengo
authors.initials: K
authors.lastname: Sakaguchi
authors.forename: Fumio
authors.initials: F
authors.lastname: Sugawara
mesh_terms: D000180:Acrylic Resins / Q000737:chemistry*; D000473:Alkanes / Q000737:chemistry* / Q000378:metabolism*; D000595:Amino Acid Sequence; D017123:Bacteriophage T7 / Q000235:genetics / Q000378:metabolism; D003001:Cloning, Molecular; D015723:Gene Library; D006801:Humans; D019169:Jurkat Cells; D007700:Kinetics; D008024:Ligands; D008969:Molecular Sequence Data; D010455:Peptides / Q000032:analysis / Q000737:chemistry / Q000378:metabolism; D010880:Piperidines / Q000737:chemistry* / Q000378:metabolism*; D011092:Polyethylene Glycols / Q000737:chemistry*; D011485:Protein Binding; D013379:Substrate Specificity; D022061:Tacrolimus Binding Protein 1A / Q000032:analysis* / Q000302:isolation & purification / Q000378:metabolism*
chemical_list: C511576:AP1497; D000180:Acrylic Resins; D000473:Alkanes; D008024:Ligands; D010455:Peptides; D010880:Piperidines; C520358:poly(acryloyl-bis(aminopropyl)polyethylene glycol); D011092:Polyethylene Glycols; D022061:Tacrolimus Binding Protein 1A