In Vivo Fast Photochemical Oxidation of Proteins Using Enhanced Multiplexing Proteomics.

Autor:	Espino JA; Department of Pharmaceutical Sciences, University of Maryland, Baltimore, Maryland 21201, United States., King CD; Department of Chemistry, Vanderbilt University, Nashville, Tennessee 37235, United States., Jones LM; Department of Pharmaceutical Sciences, University of Maryland, Baltimore, Maryland 21201, United States., Robinson RAS; Department of Chemistry, Vanderbilt University, Nashville, Tennessee 37235, United States.
Jazyk:	angličtina
Zdroj:	Analytical chemistry [Anal Chem] 2020 Jun 02; Vol. 92 (11), pp. 7596-7603. Date of Electronic Publication: 2020 May 21.
DOI:	10.1021/acs.analchem.0c00174
Abstrakt:	In vivo fast photochemical oxidation of proteins (IV-FPOP) is a hydroxyl radical protein footprinting method used to study protein structure and protein-protein interactions. Oxidatively modified proteins by IV-FPOP are analyzed by mass spectrometry (MS), and the extent of oxidation is quantified by label-free MS. Peptide oxidation changes yield useful information about protein structure, due to changes in solvent accessibility. However, the sample size necessary for animal studies requires increased sample preparation and instrument time. Here, we report the combined application of IV-FPOP and the enhanced multiplexing strategy combined precursor isotopic labeling and isobaric tagging (cPILOT) for higher-throughput analysis of oxidative modifications in C. elegans . Key differences in the performance of label-free MS and cPILOT were identified. The addition of oxygen (+16) was the most abundant modification identified among all known possible FPOP modifications. This study presents IV-FPOP coupled with enhanced multiplexing strategies such as cPILOT to increase throughput of studies seeking to examine oxidative protein modifications.
Databáze:	MEDLINE
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