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Detection of Protein Ubiquitination Sites by Peptide Enrichment and Mass Spectrometry
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Biochemistry
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JoVE Journal Biochemistry
Detection of Protein Ubiquitination Sites by Peptide Enrichment and Mass Spectrometry

Detection of Protein Ubiquitination Sites by Peptide Enrichment and Mass Spectrometry

DOI:
10.3791/59079-v

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11:54 min

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March 23, 2020

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, ,
1Proteomics Center,Erasmus University Medical Center

Chapters

  • 00:05Introduction
  • 01:16Sample Preparation
  • 03:36Offline Peptide Fractionation
  • 06:16Nanoflow LC-MS/MS
  • 07:51Data Analysis
  • 09:00Results: Detection of diGly Peptides Using Nanoflow LC-MS/MS
  • 10:51Conclusion

Summary

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We present a method for the purification, detection, and identification of diGly peptides that originate from ubiquitinated proteins from complex biological samples. The presented method is reproducible, robust, and outperforms published methods with respect to the level of depth of the ubiquitinome analysis.

Tags

Keywords: Protein UbiquitinationDiGly PeptidesMass SpectrometrySILACProtein EnrichmentLys-CTrypsin DigestionTFA Precipitation

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