Biomedical Engineering Reference
In-Depth Information
Chapter 5
Analysis of Post-translational Modifications by LC-MS/MS
Hannah Johnson and Claire E. Eyers
Abstract
Post-translational modifications are highly dynamic and known to regulate many cellular processes. Both
the site and the stoichiometry of modification of a given protein sequence can have profound effects
on the regulation of protein function. Thus, the identification of sites of post-translational modifica-
tion is crucial for fully deciphering the biological roles of any given protein. The acute regulation and
typically low stoichiometry of many post-translational modifications makes characterization of the sites
of modification challenging. Thus, the development of analytical strategies to aid the selective enrich-
ment and characterization of these species is paramount. Ongoing developments in mass spectrometry
resulting in increased speed and sensitivity of analysis mean that mass spectrometry has become the
ideal analytical tool for the qualitative and quantitative analysis of protein modifications. This chapter
provides an overview of the most popular LC-MS/MS-based strategies for the enrichment of modi-
fied peptides/proteins and mass spectrometric workflows targeted toward the analysis of specific post-
translationally modified analytes.
Key words:
Post-translational modification, mass spectrometry, LC-MS/MS, enrichment.
1.
Introduction
Post-translational modification provides a dynamic mechanism for
regulating protein function and effectively expanding the chemi-
cal diversity of functional groups and regulatory potential beyond
those defined by the standard 20 amino acid side chains. The term
post-translational modification (PTM) can refer to the addition or
the removal of a functional group from an amino acid side chain,
the modification of protein termini, the cleavage of the synthe-
sized polypeptide chain, or the covalent cross-linking between
separate protein domains. For a more comprehensive overview
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