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Probing H2O2-mediated Structural Dynamics of the Human 26S Proteasome Using Quantitative Cross-linking Mass
Clinton Yu1, Xiaorong Wang1, Alexander Scott Huszagh1
1From the ‡Department of Physiology & Biophysics, University of California, Irvine, Irvine, CA 92694.
Molecular & Cellular Proteomics : MCP
|February 7, 2019
Summary
Oxidative stress causes the 26S proteasome to weaken and reorganize. A new quantitative cross-linking mass spectrometry (QXL-MS) method reveals intermediate states before dissociation, aiding neurodegenerative disease research.
Area of Science:
- Cellular Biology
- Biochemistry
- Proteostasis
Background:
- Protein aggregation impairs cell function in neurodegenerative diseases.
- Proteasomal degradation is crucial for clearing damaged proteins.
- The 26S proteasome's response to oxidative stress is not fully understood.
Purpose of the Study:
- To investigate the structural dynamics of the 26S proteasome under oxidative stress.
- To develop a novel method for studying proteasome conformational changes.
Main Methods:
- Developed a quantitative cross-linking mass spectrometry (QXL-MS) workflow.
- Utilized SILAC labeling, affinity purification, and dual cross-linking (formaldehyde and DSSO).
- Employed multi-stage tandem mass spectrometry (MSn) for cross-link identification and quantification.
Main Results:
- Hydrogen peroxide (H2O2) treatment weakens the interaction between the 19S and 20S proteasome subcomplexes.
- Observed structural reorganizations within both the 19S and 20S subcomplexes.
- Identified potential intermediate proteasomal states preceding dissociation.
Conclusions:
- The 26S proteasome undergoes significant structural changes in response to acute oxidative stress.
- The developed QXL-MS method is effective for analyzing proteasome dynamics.
- Findings provide insights into maintaining cell viability during oxidative stress and neurodegeneration.
Keywords:
26S proteasomeOxidative stressProtein Conformation*Protein Cross-linking*Protein complex analysisProtein-Protein Interactions*Quantificationquantitative cross-linking mass spectrometrystructural dynamicsMore Related Videos
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