Constrained selected reaction monitoring: quantification of selected post-translational modifications and protein isoforms.

Journal Article

Selected reaction monitoring (SRM) is a mass spectrometry method that can target signature peptides to provide for the detection and quantitation of specific proteins in complex biological samples. When quantifying a protein, multiple peptides are generated using a specific protease such as trypsin, thereby allowing a choice of signature peptides with robust signals. In contrast, signature peptide selection can be constrained when the goal is to monitor a specific post-translational modification (PTM) or protein isoform, as the signature peptide must include the amino acid residue(s) of PTM attachment or sequence variation. This can force the selection of a signature peptide with a weak SRM response or one that is confounded by high background. In this article, we discuss steps that can be optimized to maximize peptide selection and assay performance of constrained SRM assays, including tuning instrument parameters, fragmenting product ions, using a different protease, and enriching the sample. Examples are provided for phosphorylated or citrullinated peptides and protein isoforms.

Full Text

Duke Authors

Cited Authors

  • Liu, X; Jin, Z; O'Brien, R; Bathon, J; Dietz, HC; Grote, E; Van Eyk, JE

Published Date

  • June 2013

Published In

Volume / Issue

  • 61 / 3

Start / End Page

  • 304 - 312

PubMed ID

  • 23523700

Electronic International Standard Serial Number (EISSN)

  • 1095-9130

International Standard Serial Number (ISSN)

  • 1046-2023

Digital Object Identifier (DOI)

  • 10.1016/j.ymeth.2013.03.006

Language

  • eng