Alkyne-Tag SERS Screening and Identification of Small-Molecule-Binding Sites in Protein

Jun Ando, Miwako Asanuma, Kosuke Dodo, Hiroyuki Yamakoshi, Satoshi Kawata, Katsumasa Fujita, Mikiko Sodeoka

研究成果: Article査読

44 被引用数 (Scopus)

抄録

Identification of small-molecule-binding sites in protein is important for drug discovery and analysis of protein function. Modified amino-acid residue(s) can be identified by proteolytic cleavage followed by liquid chromatography-mass spectrometry (LC-MS), but this is often hindered by the complexity of the peptide mixtures. We have developed alkyne-tag Raman screening (ATRaS) for identifying binding sites. In ATRaS, small molecules are tagged with alkyne and form covalent bond with proteins. After proteolysis and HPLC, fractions containing the labeled peptides with alkyne tags are detected by means of surface-enhanced Raman scattering (SERS) using silver nanoparticles and sent to MS/MS to identify the binding site. The use of SERS realizes high sensitivity (detection limit:100 femtomole) and reproducibility in the peptide screening. By using an automated ATRaS system, we successfully identified the inhibitor-binding site in cysteine protease cathepsin B, a potential drug target and prognostic marker for tumor metastasis. We further showed that the ATRaS system works for complex mixtures of trypsin-digested cell lysate. The ATRaS technology, which provides high molecular selectivity to LC-MS analysis, has potential to contribute in various research fields, such as drug discovery, proteomics, metabolomics and chemical biology.

本文言語English
ページ(範囲)13901-13910
ページ数10
ジャーナルJournal of the American Chemical Society
138
42
DOI
出版ステータスPublished - 2016 10 26
外部発表はい

ASJC Scopus subject areas

  • 触媒
  • 化学 (全般)
  • 生化学
  • コロイド化学および表面化学

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