Cell-permeable probe for identification and imaging of sialidases

Proc Natl Acad Sci U S A. 2013 Feb 12;110(7):2466-71. doi: 10.1073/pnas.1222183110. Epub 2013 Jan 28.

Abstract

Alkyne-hinged 3-fluorosialyl fluoride (DFSA) containing an alkyne group was shown to be a mechanism-based target-specific irreversible inhibitor of sialidases. The ester-protected analog DFSA (PDFSA) is a membrane-permeable precursor of DFSA designed to be used in living cells, and it was shown to form covalent adducts with virus, bacteria, and human sialidases. The fluorosialyl-enzyme adduct can be ligated with an azide-annexed biotin via click reaction and detected by the streptavidin-specific reporting signals. Liquid chromatography-mass spectrometry/mass spectrometry analysis on the tryptic peptide fragments indicates that the 3-fluorosialyl moiety modifies tyrosine residues of the sialidases. DFSA was used to demonstrate influenza infection and the diagnosis of the viral susceptibility to the anti-influenza drug oseltamivir acid, whereas PDFSA was used for in situ imaging of the changes of sialidase activity in live cells.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Alkynes / chemistry
  • Chromatography, Liquid
  • Click Chemistry / methods*
  • DNA Adducts / metabolism
  • Humans
  • Influenza, Human / diagnosis
  • Molecular Probe Techniques*
  • Molecular Probes / chemistry*
  • Molecular Structure
  • Neuraminidase / chemistry*
  • Neuraminidase / metabolism
  • Neuraminidase / ultrastructure*
  • Proteomics / methods
  • Streptavidin / chemistry
  • Tandem Mass Spectrometry

Substances

  • Alkynes
  • DNA Adducts
  • Molecular Probes
  • Streptavidin
  • Neuraminidase