Dataset Information

Linking post-translational modifications and protein turnover by site-resolved protein turnover (SPOT) profiling

ABSTRACT: Proteome-wide measurements of protein turnover have largely ignored the impact of post-translational modifications (PTMs). To address this gap, we employ stable isotope labelling and mass spectrometry to measure the turnover of >120,000 peptidoforms including >33,000 phosphorylated, acetylated, and ubiquitinated peptides for >9,000 native proteins. This site-resolved protein turnover (SPOT) profiling discloses global and site-specific differences in turnover associated with the presence or absence of PTMs. While causal relationships may not always be immediately apparent, we hypothesize that PTMs with diverging turnover may distinguish states of differential protein stability, structure, localization, enzymatic activity, or protein-protein interactions. We exemplify how the turnover data may facilitate insights into unknown functions of PTMs and provide a web-tool for the scientific community that allows interrogation and visualisation of all turnover data. Since the methodology is applicable to many cell types and modifications, it has substantial potential to prioritize PTMs for functional investigation in the future.

ORGANISM(S): Homo Sapiens

SUBMITTER: Julia Mergner

PROVIDER: PXD023234 | panorama | Mon Mar 14 00:00:00 GMT 2022

REPOSITORIES: PanoramaPublic

ACCESS DATA

Publications

Linking post-translational modifications and protein turnover by site-resolved protein turnover profiling.

Zecha Jana J Gabriel Wassim W Spallek Ria R Chang Yun-Chien YC Mergner Julia J Wilhelm Mathias M Bassermann Florian F Kuster Bernhard B

Nature communications 20220110 1

Proteome-wide measurements of protein turnover have largely ignored the impact of post-translational modifications (PTMs). To address this gap, we employ stable isotope labeling and mass spectrometry to measure the turnover of >120,000 peptidoforms including >33,000 phosphorylated, acetylated, and ubiquitinated peptides for >9,000 native proteins. This site-resolved protein turnover (SPOT) profiling discloses global and site-specific differences in turnover associated with the presence or absenc ...[more]

PMID: 35013197

Similar Datasets

Project description:Over the past decades, protein O-GlcNAcylation has been found to play a fundamental role in cell cycle control, metabolism, transcriptional regulation, and cellular signaling. Nevertheless, quantitative approaches to determine in vivo GlcNAc dynamics at a large-scale are still not readily available. Here, we have developed an approach to isotopically label O-GlcNAc modifications on proteins by producing 13C-labeled UDP-GlcNAc from 13C6-glucose via the hexosamine biosynthetic pathway. This metabolic labeling was combined with quantitative mass spectrometry-based proteomics to determine site-specific protein O-GlcNAcylation turnover rates. First, an efficient enrichment method for O-GlcNAc peptides was developed with the use of phenylboronic acid solid-phase extraction and anhydrous DMSO. The near stoichiometry reaction between the diol of GlcNAc and boronic acid dramatically improved the enrichment efficiency. Additionally, our kinetic model for turnover rates integrates both metabolomic and proteomic data, which increase the accuracy of the turnover rate estimation. Other advantages of this metabolic labeling method include in vivo application, direct labeling of the O-GlcNAc sites and higher confidence for site identification. Concentrating only on nuclear localized GlcNAc modified proteins, we are able to identify 159 O-GlcNAc sites on 74 proteins and determine turnover rates of 24 O-GlcNAc peptides from 21 proteins extracted from HeLa nuclei. In general, we found O-GlcNAcylation turnover rates are slower than those published for phosphorylation or acetylation. Nevertheless, the rates widely varied depending on both the protein and the residue modified. We believe this methodology can be broadly applied to reveal turnovers/dynamics of protein O-GlcNAcylation from different biological states and will provide more information on the significance of site-specific O-GlcNAcylation, enabling us to study the temporal dynamics of this critical modification in a site-specific manner for the first time.

Dataset Information

Linking post-translational modifications and protein turnover by site-resolved protein turnover (SPOT) profiling

Publications

Linking post-translational modifications and protein turnover by site-resolved protein turnover profiling.

Similar Datasets

OmicsDI is part of the ELIXIR infrastructure

Tweets