Characterization of differently processed forms of enolase 2 from Saccharomyces cerevisiae by two-dimensional gel electrophoresis and mass spectrometry

Electrophoresis. 2001 Feb;22(3):566-75. doi: 10.1002/1522-2683(200102)22:3<566::AID-ELPS566>3.0.CO;2-T.


Two-dimensional gel electrophoresis, bioinformatics, and mass spectrometry are key analysis tools in proteome analysis. The further characterization of post-translational modifications in gel-separated proteins relies fully on data obtained by mass spectrometric analysis. In this study, stress-induced changes in protein expression in Saccharomyces serevisiae were investigated. A total of eleven spots on a silver-stained two-dimensional (2-D) gel were identified by matrix-assisted laser desorption/ionization (MALDI) peptide mass mapping to represent C and/or N-terminal processed forms of enolase 2. The processing sites were determined by MALDI peptide mass mapping using a variety of proteolytic enzymes, by optimizing the sample preparation procedure and by specific labeling of all C-termini derived from in-gel digestion using a buffer containing 16O:18O (1:1). Out of eleven processed forms of enolase 2, six were fully characterized and the approximate processing sites identified for the remaining five.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Amino Acid Sequence
  • Electrophoresis, Gel, Two-Dimensional / methods*
  • Endopeptidases
  • Isotope Labeling
  • Molecular Sequence Data
  • Peptide Mapping / methods
  • Phosphopyruvate Hydratase / analysis*
  • Saccharomyces cerevisiae / enzymology*
  • Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization / methods*
  • Trypsin


  • Endopeptidases
  • Trypsin
  • Phosphopyruvate Hydratase