title: Synthesis of Grb2 SH2 Domain Proteins for Mirror-Image Screening Systems.
abstract: Grb2 is an adaptor protein that mediates cellular signal transduction. Grb2 contains an SH2 domain that interacts with phosphotyrosine-containing sequences in EGFR and other signaling molecules, and it is a promising molecular target for anticancer agents. To identify novel inhibitors of the Grb2 SH2 domain from natural products and their mirror-image isomers, screening systems using both enantiomers of a synthetic Grb2 SH2 domain protein were established. A pair of synthetic procedures for the proteins were investigated: one employed a single native chemical ligation (NCL) of two segment peptides, and the other used the N-to-C-directed NCL of three segment peptides for easier preparation. Labeling at the N-terminus or the Ala115residue of the Grb2 SH2 domain provided functional probes to detect binding to a phosphotyrosine-containing peptide. The resulting synthetic-protein-based probes were applied to bioassays, including chemical array analysis and enzyme-linked immunosorbent assays.
authors.affiliation: Graduate School of Pharmaceutical Sciences, Kyoto University , Sakyo-ku, Kyoto 606-8501, Japan.
authors.forename: Taro
authors.initials: T
authors.lastname: Noguchi
authors.affiliation: Graduate School of Pharmaceutical Sciences, Kyoto University , Sakyo-ku, Kyoto 606-8501, Japan.
authors.forename: Hiroyuki
authors.initials: H
authors.lastname: Ishiba
authors.affiliation: Chemical Biology Research Group, RIKEN Center for Sustainable Resource Science , Wako, Saitama 351-0198, Japan.
authors.forename: Kaori
authors.initials: K
authors.lastname: Honda
authors.affiliation: Chemical Biology Research Group, RIKEN Center for Sustainable Resource Science , Wako, Saitama 351-0198, Japan.
authors.forename: Yasumitsu
authors.initials: Y
authors.lastname: Kondoh
authors.affiliation: Chemical Biology Research Group, RIKEN Center for Sustainable Resource Science , Wako, Saitama 351-0198, Japan.
authors.forename: Hiroyuki
authors.identifier: 0000-0001-7400-8388
authors.initials: H
authors.lastname: Osada
authors.affiliation: Graduate School of Pharmaceutical Sciences, Kyoto University , Sakyo-ku, Kyoto 606-8501, Japan.
authors.forename: Hiroaki
authors.initials: H
authors.lastname: Ohno
authors.affiliation: Graduate School of Pharmaceutical Sciences, Kyoto University , Sakyo-ku, Kyoto 606-8501, Japan.
authors.forename: Nobutaka
authors.initials: N
authors.lastname: Fujii
authors.affiliation: Graduate School of Pharmaceutical Sciences, Kyoto University , Sakyo-ku, Kyoto 606-8501, Japan.
authors.forename: Shinya
authors.identifier: 0000-0002-2833-2539
authors.initials: S
authors.lastname: Oishi
mesh_terms: D000595:Amino Acid Sequence; D055808:Drug Discovery / Q000379:methods*; D004797:Enzyme-Linked Immunosorbent Assay; D051380:GRB2 Adaptor Protein / Q000037:antagonists & inhibitors / Q000138:chemical synthesis* / Q000737:chemistry / Q000378:metabolism; D006801:Humans; D008958:Models, Molecular; D010455:Peptides / Q000737:chemistry / Q000494:pharmacology; D018909:src Homology Domains / Q000187:drug effects*
chemical_list: D051380:GRB2 Adaptor Protein; D010455:Peptides