An in vivo assay for the identification of target proteases which cleave membrane-associated substrates

FEBS Lett. 1999 Dec 17;463(3):245-9. doi: 10.1016/s0014-5793(99)01627-0.

Abstract

Proteases not only play a fundamental role in numerous physiological processes, but are also involved in several human diseases including Alzheimer's disease (AD). A key protease implicated in AD is the so far unidentified gamma-secretase, which cleaves the membrane-bound beta-amyloid precursor protein (betaAPP) at the C-terminus of its amyloid domain within the membrane to release the neurotoxic amyloid beta-peptide. In order to allow the isolation of proteases, which specifically cleave membrane-bound substrates within or in the vicinity of a transmembrane domain, we developed a reporter gene assay in Saccharomyces cerevisiae. This assay may allow the identification of genes encoding target proteases that specifically cleave membrane bound substrates by transforming expression libraries.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Amyloid Precursor Protein Secretases
  • Aspartic Acid Endopeptidases
  • Binding Sites
  • Caspase 3
  • Caspases / genetics
  • DNA-Binding Proteins
  • Endopeptidases / analysis*
  • Endopeptidases / genetics
  • Endopeptidases / metabolism
  • Fungal Proteins / genetics
  • Genes, Reporter
  • Membrane Proteins / analysis
  • Membrane Proteins / metabolism*
  • Plasmids
  • Proteins / genetics
  • Receptors, Notch
  • Recombinant Fusion Proteins / biosynthesis
  • Recombinant Fusion Proteins / genetics
  • Saccharomyces cerevisiae / genetics
  • Saccharomyces cerevisiae / metabolism
  • Saccharomyces cerevisiae Proteins*
  • Substrate Specificity
  • Transcription Factors / genetics

Substances

  • DNA-Binding Proteins
  • Fungal Proteins
  • GAL4 protein, S cerevisiae
  • Membrane Proteins
  • Proteins
  • Receptors, Notch
  • Recombinant Fusion Proteins
  • Saccharomyces cerevisiae Proteins
  • Transcription Factors
  • Amyloid Precursor Protein Secretases
  • Endopeptidases
  • CASP3 protein, human
  • Caspase 3
  • Caspases
  • Aspartic Acid Endopeptidases
  • BACE1 protein, human