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Versatile assays for high throughput screening for activators or inhibitors of intracellular proteases and their cellular regulators
- Source :
- PLoS ONE, PLoS ONE, Vol 4, Iss 10, p e7655 (2009)
- Publication Year :
- 2009
-
Abstract
- BACKGROUND: Intracellular proteases constitute a class of promising drug discovery targets. Methods for high throughput screening against these targets are generally limited to in vitro biochemical assays that can suffer many technical limitations, as well as failing to capture the biological context of proteases within the cellular pathways that lead to their activation. METHODS #ENTITYSTARTX00026; FINDINGS: We describe here a versatile system for reconstituting protease activation networks in yeast and assaying the activity of these pathways using a cleavable transcription factor substrate in conjunction with reporter gene read-outs. The utility of these versatile assay components and their application for screening strategies was validated for all ten human Caspases, a family of intracellular proteases involved in cell death and inflammation, including implementation of assays for high throughput screening (HTS) of chemical libraries and functional screening of cDNA libraries. The versatility of the technology was also demonstrated for human autophagins, cysteine proteases involved in autophagy. CONCLUSIONS: Altogether, the yeast-based systems described here for monitoring activity of ectopically expressed mammalian proteases provide a fascile platform for functional genomics and chemical library screening.<br />PloS one, 4(10), e7655; 2009
- Subjects :
- Proteases
Transcription, Genetic
medicine.medical_treatment
High-throughput screening
Chemistry, Pharmaceutical
Drug Evaluation, Preclinical
lcsh:Medicine
Computational biology
Biology
Chemical library
Small Molecule Libraries
03 medical and health sciences
chemistry.chemical_compound
Structure-Activity Relationship
0302 clinical medicine
Genes, Reporter
Peptide Library
Chemical Biology
medicine
Combinatorial Chemistry Techniques
Humans
Protease Inhibitors
lcsh:Science
030304 developmental biology
Gene Library
Inflammation
0303 health sciences
Reporter gene
Caspase 8
Multidisciplinary
Protease
Drug discovery
lcsh:R
Cell Biology
Molecular biology
chemistry
030220 oncology & carcinogenesis
Drug Design
lcsh:Q
Functional genomics
Research Article
Biotechnology
Peptide Hydrolases
Subjects
Details
- Language :
- English
- ISSN :
- 19326203
- Volume :
- 4
- Issue :
- 10
- Database :
- OpenAIRE
- Journal :
- PloS one
- Accession number :
- edsair.doi.dedup.....f1b6725a3db4b2df2cc9b02343e8b3b6