Synthesis of Grb2 SH2 Domain Proteins for Mirror-Image Screening Systems

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• 作者：Taro Noguchi ; Hiroyuki Ishiba ; Kaori Honda ; Yasumitsu Kondoh ; Hiroyuki OsadaHiroaki Ohno ; Nobutaka Fujii ; Shinya Oishi
• 刊名：Bioconjugate Chemistry
• 出版年：2017
• 出版时间：February 15, 2017
• 年：2017
• 卷：28
• 期：2
• 页码：609-619
• 全文大小：820K
• ISSN：1520-4812

文摘

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 Ala¹¹⁵ residue 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.