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Protocol for proteomic profiling of hypusinated protein candidates using a clickable spermidine probe
Tian Zhang1, Jianlong Li1, Xinbo Hu2
1Interdisciplinary Research Center on Biology and Chemistry, Shanghai Institute of Organic Chemistry, Chinese Academy of Sciences, Shanghai, China; University of Chinese Academy of Sciences, Beijing, China.
STAR Protocols
|August 10, 2026
Summary
This study introduces a new method for identifying hypusinated proteins. The protocol uses a special probe to detect proteins modified by deoxyhypusine synthase (DHPS) and deoxyhypusine hydroxylase (DOHH).
Area of Science:
- Biochemistry
- Proteomics
- Molecular Biology
Background:
- Hypusination is a unique posttranslational modification involving deoxyhypusine synthase (DHPS) and deoxyhypusine hydroxylase (DOHH).
- Identifying hypusinated proteins is crucial for understanding their biological roles.
Purpose of the Study:
- To develop a proteome-wide protocol for identifying candidate hypusinated proteins.
- To enable comprehensive analysis of hypusination in cellular systems.
Main Methods:
- Synthesis of a clickable alkynyl-spermidine probe.
- DHPS-dependent metabolic labeling and click chemistry-mediated biotinylation.
- Streptavidin-based enrichment followed by mass spectrometry analysis.
Main Results:
- A robust protocol for the proteome-wide identification of hypusinated proteins was established.
- The method allows for sensitive detection and characterization of hypusinated targets.
Conclusions:
- This protocol provides a powerful tool for studying hypusination.
- It facilitates further research into the functions and implications of hypusinated proteins.
