SimPLIT: Simplified Sample Preparation for Large-Scale Isobaric Tagging Proteomics.

Content Provider	Europe PMC
Author	Sialana, FernandoJ. Roumeliotis, Theodoros I. Bouguenina, Habib Chan Wah Hak, Laura Wang, Hannah Caldwell, John Collins, Ian Chopra, Rajesh Choudhary, Jyoti S.
Copyright Year	2022
Abstract	Large scale proteomic profiling of cell lines can revealmolecularsignatures attributed to variable genotypes or induced perturbations,enabling proteogenomic associations and elucidation of pharmacologicalmechanisms of action. Although isobaric labeling has increased thethroughput of proteomic analysis, the commonly used sample preparationworkflows often require time-consuming steps and costly consumables,limiting their suitability for large scale studies. Here, we presenta simplified and cost-effective one-pot reaction workflow in a 96-wellplate format (SimPLIT) that minimizes processing steps and demonstratesimproved reproducibility compared to alternative approaches. The workflowis based on a sodium deoxycholate lysis buffer and a single detergentcleanup step after peptide labeling, followed by quick off-line fractionationand MS2 analysis. We showcase the applicability of the workflow ina panel of colorectal cancer cell lines and by performing target discoveryfor a set of molecular glue degraders in different cell lines, ina 96-sample assay. Using this workflow, we report frequently dysregulatedproteins in colorectal cancer cells and uncover cell-dependent proteindegradation profiles of seven cereblon E3 ligase modulators (CRL4CRBN). Overall, SimPLIT is a robust method that can be easilyimplemented in any proteomics laboratory for medium-to-large scaleTMT-based studies for deep profiling of cell lines.
Related Links	https://europepmc.org/backend/ptpmcrender.fcgi?accid=PMC9361352&blobtype=pdf
ISSN	15353893
Journal	Journal of Proteome Research [J Proteome Res]
Volume Number	21
DOI	10.1021/acs.jproteome.2c00092
PubMed Central reference number	PMC9361352
Issue Number	8
PubMed reference number	35848491
e-ISSN	15353907
Language	English
Publisher	American Chemical Society
Publisher Date	2022-07-18
Access Restriction	Open
Rights License	Permits the broadest form of re-use including for commercial purposes, provided that author attribution and integrity are maintained (https://creativecommons.org/licenses/by/4.0/). © 2022 The Authors. Published by American Chemical Society
Subject Keyword	isobaric labeling TMTpro cancer cell lines targeted protein degradation IMiDs CELMoDs
Content Type	Text
Resource Type	Article
Subject	Chemistry Biochemistry