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Updated: Aug 12, 2026
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Profiling Ubiquitin and Ubiquitin-like Dependent Post-translational Modifications and Identification of Significant Alterations
Published on: November 7, 2019
A ubiquitin-based tagging system for controlled modulation of protein stability
J H Stack1, M Whitney, S M Rodems
1Aurora Biosciences Corporation, 11010 N. Torreyana Road, San Diego, CA 92121, USA.
Nature Biotechnology
|December 2, 2000
Summary
Researchers created a system to control protein stability in mammalian cells using destabilization domains. This system enables tunable protein half-life and steady-state levels for biotechnology applications.
Area of Science:
- Biotechnology
- Molecular Biology
- Cell Biology
Background:
- Controllable protein expression is crucial for biotechnology.
- Protein stability, or half-life, dictates intracellular protein concentration.
- Existing methods for controlling protein stability are limited.
Purpose of the Study:
- To develop a versatile and reliable system for producing short half-life proteins in mammalian cells.
- To enable tunable control over protein stability and steady-state levels.
- To establish a high-throughput screening assay for proteasome activity.
Main Methods:
- Engineered destabilization domains using varying numbers of a mutant ubiquitin form (UbG76V).
- Fused destabilization domains to heterologous reporter proteins and cellular proteins.
- Assessed protein half-life and steady-state levels in mammalian cells.
- Utilized a destabilized beta-lactamase reporter for proteasome activity screening.
Main Results:
- Increasing UbG76V moieties in destabilization domains resulted in a graded decrease in protein half-life.
- Steady-state protein levels were reduced proportionally to the decrease in half-life.
- Cells expressing destabilized beta-lactamase served as a robust, HTS-compatible assay for proteasome activity.
Conclusions:
- The developed system provides a versatile method for controlling protein half-life in mammalian cells.
- This system allows for tunable regulation of intracellular protein concentrations.
- The destabilized reporter system is effective for high-throughput screening of proteasome activity.
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02:04Covalently Linked Protein Regulators
Proteins can undergo many types of post-translational modifications, often in response to changes in their environment. These modifications play an important role in the function and stability of these proteins. Covalently linked molecules include functional groups, such as methyl, acetyl, and phosphate groups, and also small proteins, such as ubiquitin. There are around 200 different types of covalent regulators that have been identified.
These groups modify specific amino acids in a protein.
These groups modify specific amino acids in a protein.
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