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Keywords:

  • GPSP;
  • metabolic labelling;
  • protein degradation;
  • proteolysis;
  • proteostasis;
  • pulse-chase;
  • SILAC

Abstract

Protein degradation is a critical factor in controlling cellular protein abundance. Here, we compare classical methods for determining protein degradation rates to a novel GFP (green fluorescent protein) fusion protein based method that assesses the intrinsic stability of cloned cDNA library products by flow cytometry [Yen et al. (2008) Science 322, 918]. While no method is perfect, we conclude that chimeric gene reporter approaches, though powerful, should be applied cautiously, due principally to GFP (or other reporter tag) interference with protein organelle targeting or incorporation into macromolecular assemblies, both of which cause spuriously high degradation rates.