Mass spectrometric analysis of PTM dynamics using stable isotope labeled metabolic precursors in cell culture

Pijkeren, Alienke van; Bischoff, Rainer; Kwiatkowski, Marcel

doi:10.1039/c9an01258c

Cited by 5 publications

(9 citation statements)

References 175 publications

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“…As acetyl-CoA is also the substrate for histone acetylations, natural occurring 12 C-acetylation of lysines are replaced with 13 Clabelled acetyl-groups causing a characteristic mass shift of the peptide in high-resolution MS spectra. The timedependent replacement of 12 C-acetylation can be used to determine histone acetylation dynamics [8,9,13,14].…”

Section: Resultsmentioning

confidence: 99%

Natural isotope correction improves analysis of protein modification dynamics

et al. 2021

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Stable isotope labelling in combination with high-resolution mass spectrometry approaches are increasingly used to analyze both metabolite and protein modification dynamics. To enable correct estimation of the resulting dynamics, it is critical to correct the measured values for naturally occurring stable isotopes, a process commonly called isotopologue correction or deconvolution. While the importance of isotopologue correction is well recognized in metabolomics, it has received far less attention in proteomics approaches. Although several tools exist that enable isotopologue correction of mass spectrometry data, the majority is tailored for the analysis of low molecular weight metabolites. We here present PICor which has been developed for isotopologue correction of complex isotope labelling experiments in proteomics or metabolomics and demonstrate the importance of appropriate correction for accurate determination of protein modifications dynamics, using histone acetylation as an example.

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Section: Resultsmentioning

confidence: 99%

Natural isotope correction improves analysis of protein modification dynamics

et al. 2021

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“…As acetyl-CoA is also the substrate for histone acetylations, natural occuring 12 C-acetylation of lysines are replaced with 13 C-labelled acetyl-groups causing a characteristic mass shift of the peptide in high resolution MS-spectra. The time dependent replacement of 12 C-acetylation can be used to determine histone acetylation dynamics [1][2][3]6 .…”

Section: Resultsmentioning

confidence: 99%

“…While it is well known that protein phosphorylations are highly dynamic and result from a complex interplay between protein kinases and phosphatases, the dynamics of other protein modifications such as methylation and acetylation have been less well investigated. The availability of stable isotope labelled compounds and high resolution mass spectrometry has, however, more recently paved the way to analyse the dynamics of protein methylation, acetylation, phosphorylation and glycosylation [1][2][3][4][5] .…”

Section: Introductionmentioning

confidence: 99%

Natural isotope correction improves analysis of protein modification dynamics

Dietze

Pijkeren

Egger

et al. 2020

Preprint

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Stable isotope labelling in combination with high resolution mass spectrometry approaches are increasingly used to analyse both metabolite and protein modification dynamics. To enable correct estimation of the resulting dynamics it is critical to correct the measured values for naturally occurring stable isotopes, a process commonly called isotopologue correction or deconvolution. While the importance of isotopologue correction is well recognized in metabolomics, it has received far less attention in proteomics approaches. Although several tools exist that enable isotopologue correction of mass spectrometry data, none of them is universally applicable for all potential experimental approaches. We here present PICor which has been streamlined for multiple isotope labelling isotopologue correction in proteomics or metabolomics approaches. We demonstrate the importance for accurate measurement of the dynamics of protein modifications, such as histone acetylation.

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“…They do not allow us to monitor site-specific acetylation dynamics, which is critical to follow the regulation, turnover, and interplay of histone modifications. Stable isotope labeled metabolic precursors can be used to study dynamics of proteins and posttranslational modifications [14] . To study histone acetylation dynamics, Evertts et al [15] and Zheng et al [16] introduced metabolic labeling by uniformly 13 C labeled glucose ([U-13C]-Glc) as a metabolic precursor.…”

Section: Introductionmentioning

confidence: 99%

Combined Metabolic and Chemical (CoMetChem) Labeling Using Stable Isotopes – a Strategy to Reveal Site-Specific Histone Acetylation and Deacetylation Rates by LC-MS

Pijkeren¹,

Dietze²,

Brotons³

et al. 2021

Preprint

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Histone acetylation is an important, reversible post-translational protein modification and a hallmark of epigenetic regulation. However, little is known about the dynamics of this process, due to the lack of analytical methods that can capture site-specific acetylation and deacetylation reactions. We present a new approach that combines metabolic and chemical labeling (CoMetChem) using uniformly 13C-labeled glucose and stable isotope labeled acetic anhydride. Thereby, chemically equivalent, fully acetylated histone species are generated enabling accurate relative quantification of site-specific lysine acetylation in tryptic peptides using high-resolution mass spectrometry. We show that CoMetChem enables site-specific quantification of the incorporation or loss of lysine acetylation over time, allowing the determination of reaction rates for acetylation and deacetylation. Thus, the CoMetChem methodology provides a comprehensive description of site-specific acetylation dynamics. <br>

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Mass spectrometric analysis of PTM dynamics using stable isotope labeled metabolic precursors in cell culture

Abstract: Metabolic stable isotope labeled precursors allow the study of PTM dynamics by mass spectrometry providing essential information to understand biological processes.

Cited by 5 publications

References 175 publications

Natural isotope correction improves analysis of protein modification dynamics

Natural isotope correction improves analysis of protein modification dynamics

Natural isotope correction improves analysis of protein modification dynamics

Combined Metabolic and Chemical (CoMetChem) Labeling Using Stable Isotopes – a Strategy to Reveal Site-Specific Histone Acetylation and Deacetylation Rates by LC-MS

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