title: Protein turnover measurement using selected reaction monitoring-mass spectrometry (SRM-MS).
abstract: Protein turnover represents an important mechanism in the functioning of cells, with deregulated synthesis and degradation of proteins implicated in many diseased states. Therefore, proteomics strategies to measure turnover rates with high confidence are of vital importance to understanding many biological processes. In this study, the more widely used approach of non-targeted precursor ion signal intensity (MS1) quantification is compared with selected reaction monitoring (SRM), a data acquisition strategy that records data for specific peptides, to determine if improved quantitative data would be obtained using a targeted quantification approach. Using mouse liver as a model system, turnover measurement of four tricarboxylic acid cycle proteins was performed using both MS1 and SRM quantification strategies. SRM outperformed MS1 in terms of sensitivity and selectivity of measurement, allowing more confident determination of protein turnover rates. SRM data are acquired using cheaper and more widely available tandem quadrupole mass spectrometers, making the approach accessible to a larger number of researchers than MS1 quantification, which is best performed on high mass resolution instruments. SRM acquisition is ideally suited to focused studies where the turnover of tens of proteins is measured, making it applicable in determining the dynamics of proteins complexes and complete metabolic pathways.This article is part of the themed issue 'Quantitative mass spectrometry'.
authors.affiliation: Centre for Proteome Research, Department of Biochemistry, Institute of Integrative Biology, University of Liverpool, Crown Street, Liverpool L69 7ZB, UK stephen.holman@liverpool.ac.uk.
authors.forename: Stephen W
authors.initials: SW
authors.lastname: Holman
authors.affiliation: Cellular and Molecular Physiology, Institute of Translational Medicine, University of Liverpool, Crown Street, Liverpool L69 3BX, UK.
authors.forename: Dean E
authors.initials: DE
authors.lastname: Hammond
authors.affiliation: Centre for Proteome Research, Department of Biochemistry, Institute of Integrative Biology, University of Liverpool, Crown Street, Liverpool L69 7ZB, UK.
authors.forename: Deborah M
authors.initials: DM
authors.lastname: Simpson
authors.affiliation: Mammalian Behaviour and Evolution Group, Department of Evolution, Ecology and Behaviour, Institute of Integrative Biology, University of Liverpool, Leahurst Campus, Neston CH64 7TE, UK.
authors.forename: John
authors.initials: J
authors.lastname: Waters
authors.affiliation: Mammalian Behaviour and Evolution Group, Department of Evolution, Ecology and Behaviour, Institute of Integrative Biology, University of Liverpool, Leahurst Campus, Neston CH64 7TE, UK.
authors.forename: Jane L
authors.initials: JL
authors.lastname: Hurst
authors.affiliation: Centre for Proteome Research, Department of Biochemistry, Institute of Integrative Biology, University of Liverpool, Crown Street, Liverpool L69 7ZB, UK r.beynon@liverpool.ac.uk.
authors.forename: Robert J
authors.identifier: http://orcid.org/0000-0003-0857-495X
authors.initials: RJ
authors.lastname: Beynon
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D013058:Mass Spectrometry / Q000379:methods*; D051379:Mice; D011506:Proteins / Q000737:chemistry / Q000378:metabolism*; D040901:Proteomics / Q000379:methods*
chemical_list: D011506:Proteins
keywords: MS1; SRM; mass spectrometry; protein turnover; proteomics; quantification