An in vivo assay for the identification of target proteases which cleave membrane-associated substrates.
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.
Steiner
H
H
Central Institute of Mental Health, Department of Molecular Biology, J5, 68159, Mannheim, Germany.
Pesold
B
B
Haass
C
C
D053829:Amyloid Precursor Protein Secretases; D016282:Aspartic Acid Endopeptidases; D001665:Binding Sites; D053148:Caspase 3; D020169:Caspases / Q000235:genetics; D004268:DNA-Binding Proteins; D010450:Endopeptidases / Q000032:analysis* / Q000235:genetics / Q000378:metabolism; D005656:Fungal Proteins / Q000235:genetics; D017930:Genes, Reporter; D008565:Membrane Proteins / Q000032:analysis / Q000378:metabolism*; D010957:Plasmids; D011506:Proteins / Q000235:genetics; D051880:Receptors, Notch; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis / Q000235:genetics; D012441:Saccharomyces cerevisiae / Q000235:genetics / Q000378:metabolism; D029701:Saccharomyces cerevisiae Proteins; D013379:Substrate Specificity; D014157:Transcription Factors / Q000235:genetics
D004268:DNA-Binding Proteins; D005656:Fungal Proteins; C052130:GAL4 protein, S cerevisiae; D008565:Membrane Proteins; D011506:Proteins; D051880:Receptors, Notch; D011993:Recombinant Fusion Proteins; D029701:Saccharomyces cerevisiae Proteins; D014157:Transcription Factors; D053829:Amyloid Precursor Protein Secretases; D010450:Endopeptidases; C505407:CASP3 protein, human; D053148:Caspase 3; D020169:Caspases; D016282:Aspartic Acid Endopeptidases; C509280:BACE1 protein, human