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Updated: Aug 22, 2026
Biosensor-based High Throughput Biopanning and Bioinformatics Analysis Strategy for the Global Validation of Drug-protein Interactions
Published on: December 1, 2020
Profiling treatment-specific post-translational modifications in a complex proteome with subtractive substrate phage
Angela Tenzer1, Barbara Hofstetter, Christelle Sauser
1Department of Radiation Oncology, University Hospital Zurich, Zurich, Switzerland.
Abstract:
Proteolytic activation of zymogens or controlled degradation of inhibitory factors is part of a major regulatory system on the post-translational level to regulate treatment induced cellular stress responses. The identification of differential activity based substrates is thus of high interest to prioritize and validate candidate targets for drug discovery. Here we present a novel subtractive substrate phage display screening method for the selection of treatment induced post-translational peptide modifications in complex proteomes. We investigated this approach with tumor cells in response to a protease activating anticancer treatment modality using subtractive and iterative screening of cellular extracts derived from control and treated cells. Specific phage were identified that served as substrates for proteolytic activities in response to treatment related activity changes and could be distinguished from substrates for unspecific proteolytic background activities. Novel, selected peptide substrates were investigated in vitro and in vivo and showed high substrate specificity and functional biological significance.
Insights
Researchers developed a new method to identify treatment-induced protein modifications. This technique aids in discovering new drug targets by finding specific enzyme substrates in complex biological samples.
Area of Science:
- Biochemistry
- Molecular Biology
- Proteomics
Background:
- Post-translational modifications regulate cellular stress responses.
- Identifying substrates of enzymes with altered activity is crucial for drug discovery.
- Targeting treatment-induced modifications offers a novel therapeutic strategy.
Purpose of the Study:
- To present a novel subtractive substrate phage display screening method.
- To select treatment-induced post-translational peptide modifications in complex proteomes.
- To identify and validate novel peptide substrates for drug discovery.
Main Methods:
- Subtractive and iterative phage display screening of cellular extracts.
- Utilized tumor cells treated with a protease-activating anticancer therapy.
- Distinguished specific substrates from background proteolytic activity.
Main Results:
- Identified specific phages acting as substrates for treatment-induced proteolytic activities.
- Successfully differentiated treatment-specific substrates from non-specific ones.
- Validated novel peptide substrates in vitro and in vivo, demonstrating specificity and biological significance.
Conclusions:
- The developed subtractive substrate phage display is effective for identifying treatment-induced proteolytic modifications.
- Novel peptide substrates identified have high specificity and biological relevance.
- This method facilitates the prioritization and validation of drug discovery targets.
