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Simultaneous Affinity Enrichment of Two Post-Translational Modifications for Quantification and Site Localization
Published on: February 27, 2020
Multiple neutral loss monitoring (MNM): a multiplexed method for post-translational modification screening
Michael D Hoffman1, Matthew J Sniatynski, Jason C Rogalski
1The Biomedical Research Centre, University of British Columbia, Vancouver, British Columbia, Canada.
Journal of the American Society for Mass Spectrometry
|January 31, 2006
Summary
A new mass spectrometry method, multiple neutral loss monitoring (MNM), comprehensively screens for post-translational modifications (PTMs) without prior specification. This advance allows for broader protein characterization by detecting all PTMs that fragment as neutral losses.
Area of Science:
- Proteomics and Mass Spectrometry
- Biochemistry and Molecular Biology
Background:
- Post-translational modifications (PTMs) are crucial for protein function and activity.
- Current mass spectrometry methods require pre-specification of PTMs, limiting comprehensive analysis.
- This necessitates multiple analyses for extensive protein characterization.
Purpose of the Study:
- To develop a novel mass spectrometry scan strategy for comprehensive screening of PTMs.
- To enable the detection of all PTMs that fragment as neutral losses.
- To overcome limitations of current methods requiring specific PTM identification prior to analysis.
Main Methods:
- Development of the multiple neutral loss monitoring (MNM) scan strategy.
- Determination of MNM method parameters using product ion scans and collision energies.
- Optimization of collision energies for different PTMs and peptide charge states using a collision energy gradient.
- Application of autocorrelation and convolution mapping for modification and peptide identification.
- Validation using a BSA digest mixture containing four modified peptides.
Main Results:
- The MNM method was optimized by establishing neutral loss energy profiles and optimal collision energies (OCE) for various modifications.
- A collision energy gradient was implemented in the MNM scan due to the dependence of OCE on modification type and peptide charge state.
- Autocorrelation and convolution mapping successfully identified modification types and associated peptides.
- The MNM scan was successfully applied to identify multiple modified peptides in a complex biological sample (BSA digest).
Conclusions:
- The novel MNM scan strategy allows for comprehensive screening of PTMs that fragment as neutral losses.
- This technique significantly enhances the scope of protein characterization in mass spectrometry-based proteomics.
- Implementation of MNM will facilitate a more complete understanding of protein function and regulation through extensive PTM analysis.
