Rapid and ultra-sensitive determination of enzyme activities using surface-enhanced resonance Raman scattering

B.D. Moore, L. Stevenson, A. Watt, S. Flitsch, N.J. Turner, C. Cassidy, D. Graham

Research output: Contribution to journalArticle

165 Citations (Scopus)

Abstract

Measurement of enzyme activity and selectivity at in vivo concentrations is highly desirable in a range of fields including diagnostics, functional proteomics and directed evolution. Here we demonstrate how surface-enhanced resonance Raman scattering (SERRS), measured using silver nanoparticles, can be used to detect the activity of hydrolases at ultra-low levels. This approach was made possible by designing 'masked' enzyme substrates that are initially completely undetected by SERRS. Turnover of the substrate by the enzyme leads to the release of a surface targeting dye, and intense SERRS signals proportional to enzyme activity are generated. The method was used to rapidly screen the relative activities and enantioselectivities of fourteen enzymes including examples of lipases, esterases and proteases. In the current format the sensitivity of the technique is sufficient to detect 500 enzyme molecules, which offers the potential to detect multiple enzyme activities simultaneously and at levels found within single cells.
Original languageEnglish
Pages (from-to)1133-1138
Number of pages5
JournalNature Biotechnology
Volume22
Issue number9
DOIs
Publication statusPublished - Sep 2004

Keywords

  • enantioselective hydrolases
  • ph indicators
  • chemistry
  • catalysts
  • silver
  • serrs
  • benzotriazole
  • spectroscopy
  • resolution
  • libraries

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