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Title: Plasma-Generated OH Radical Production for Analyzing Three-Dimensional Structure in Protein Therapeutics

Abstract

Protein three-dimensional structure dynamically changes in solution depending on the presence of ligands and interacting proteins. Methods for detecting these changes in protein conformation include ‘protein footprinting,’ using mass spectrometry. We describe herein a new technique, PLIMB (Plasma Induced Modification of Biomolecules), that generates µs bursts of hydroxyl radicals from water, to measure changes in protein structure via altered solvent accessibility of amino acid side chains. PLIMB was first benchmarked with model compounds, and then applied to a biological problem, i.e., ligand (EGF) induced changes in the conformation of the external (ecto) domain of Epidermal Growth Factor Receptor (EGFR). Regions in which oxidation decreased upon adding EGF fall along the dimerization interface, consistent with models derived from crystal structures. Furthermore these results demonstrate that plasma-generated hydroxyl radicals from water can be used to map protein conformational changes, and provide a readily accessible means of studying protein structure in solution.

Authors:
 [1];  [1];  [1];  [1];  [1];  [1]
  1. Univ. of Wisconsin-Madison, Madison, WI (United States)
Publication Date:
Research Org.:
Univ. of Wisconsin, Madison, WI (United States)
Sponsoring Org.:
USDOE
OSTI Identifier:
1500095
Grant/Contract Number:  
FG02-88ER13938
Resource Type:
Accepted Manuscript
Journal Name:
Scientific Reports
Additional Journal Information:
Journal Volume: 7; Journal Issue: 1; Journal ID: ISSN 2045-2322
Publisher:
Nature Publishing Group
Country of Publication:
United States
Language:
English
Subject:
59 BASIC BIOLOGICAL SCIENCES

Citation Formats

Minkoff, Benjamin B., Blatz, Joshua M., Choudhury, Faraz A., Benjamin, Daniel, Shohet, J. Leon, and Sussman, Michael R. Plasma-Generated OH Radical Production for Analyzing Three-Dimensional Structure in Protein Therapeutics. United States: N. p., 2017. Web. doi:10.1038/s41598-017-13371-7.
Minkoff, Benjamin B., Blatz, Joshua M., Choudhury, Faraz A., Benjamin, Daniel, Shohet, J. Leon, & Sussman, Michael R. Plasma-Generated OH Radical Production for Analyzing Three-Dimensional Structure in Protein Therapeutics. United States. doi:10.1038/s41598-017-13371-7.
Minkoff, Benjamin B., Blatz, Joshua M., Choudhury, Faraz A., Benjamin, Daniel, Shohet, J. Leon, and Sussman, Michael R. Wed . "Plasma-Generated OH Radical Production for Analyzing Three-Dimensional Structure in Protein Therapeutics". United States. doi:10.1038/s41598-017-13371-7. https://www.osti.gov/servlets/purl/1500095.
@article{osti_1500095,
title = {Plasma-Generated OH Radical Production for Analyzing Three-Dimensional Structure in Protein Therapeutics},
author = {Minkoff, Benjamin B. and Blatz, Joshua M. and Choudhury, Faraz A. and Benjamin, Daniel and Shohet, J. Leon and Sussman, Michael R.},
abstractNote = {Protein three-dimensional structure dynamically changes in solution depending on the presence of ligands and interacting proteins. Methods for detecting these changes in protein conformation include ‘protein footprinting,’ using mass spectrometry. We describe herein a new technique, PLIMB (Plasma Induced Modification of Biomolecules), that generates µs bursts of hydroxyl radicals from water, to measure changes in protein structure via altered solvent accessibility of amino acid side chains. PLIMB was first benchmarked with model compounds, and then applied to a biological problem, i.e., ligand (EGF) induced changes in the conformation of the external (ecto) domain of Epidermal Growth Factor Receptor (EGFR). Regions in which oxidation decreased upon adding EGF fall along the dimerization interface, consistent with models derived from crystal structures. Furthermore these results demonstrate that plasma-generated hydroxyl radicals from water can be used to map protein conformational changes, and provide a readily accessible means of studying protein structure in solution.},
doi = {10.1038/s41598-017-13371-7},
journal = {Scientific Reports},
number = 1,
volume = 7,
place = {United States},
year = {2017},
month = {10}
}

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Cited by: 5 works
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Figures / Tables:

Figure 1 Figure 1: Microsecond bursts of radicals modify free methionine and BSA in a dose dependent fashion. (A) Left, schematic of sample and instrument. Right, photograph demonstrating treatment in progress. (B) Graph demonstrating μs breakdowns to produce μs bursts of hydroxyl radicals. (C) Production of oxidized methionine with increasing plasma treatmentmore » time. For each data point, n = 3 and error bars are ± standard deviation. (D) Modification observed on native (above) and digested (below) BSA with increasing plasma treatment times. Regions shown for comparison have been selected; full data in Fig. S3. For each bar, n = 3 and error bars are ± standard deviation.« less

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