BioRxiv
bioRxiv Subject Collection: Systems Biology
This feed contains articles for bioRxiv Subject Collection "Systems Biology"
An integrated workflow for quantitative analysis of the newly synthesized proteome
The analysis of proteins that are newly synthesized upon a cellular perturbation can provide detailed insight in the proteomic response that is elicited by specific cues. This can be investigated by pulse-labeling of cells with clickable and stable-isotope-coded amino acids for enrichment and mass spectrometric characterization of newly synthesized proteins (NSPs), however convoluted protocols prohibit their routine application. Here we optimized multiple steps in sample preparation, mass spectrometry and data analysis, and integrated them in a semi-automated workflow for the quantitative analysis of the newly synthesized proteome (QuaNPA). Reduced input requirements and data-independent acquisition (DIA) enabled analysis of triple-SILAC-labeled NSP samples, with enhanced throughput while featuring high quantitative accuracy. We applied QuaNPA to investigate the time-resolved cellular response to interferon-gamma (IFNg), observing rapid induction of known and novel targets 2h after IFNg treatment. QuaNPA provides a powerful approach for large-scale investigation of NSPs to gain insight in complex cellular processes.
Bortecen, T., Mueller, T., Krijgsveld, J.
March 4, 2023
http://biorxiv.org/cgi/content/short/2023.03.03.530942v1?rss=1