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A Methodology for Simultaneous Fluorogenic Derivatization and Boronate Affinity Enrichment of 3-Nitrotyrosine Containing Peptides
dc.contributor.author | Dremina, Elena S. | |
dc.contributor.author | Li, Xiaobao | |
dc.contributor.author | Galeva, Nadezhda A. | |
dc.contributor.author | Sharov, Victor S. | |
dc.contributor.author | Stobaugh, John F. | |
dc.contributor.author | Schoneich, Christian | |
dc.date.accessioned | 2017-01-12T19:22:48Z | |
dc.date.available | 2017-01-12T19:22:48Z | |
dc.date.issued | 2012-11-15 | |
dc.identifier.citation | Dremina, Elena S., Xiaobao Li, Nadezhda A. Galeva, Victor S. Sharov, John F. Stobaugh, and Christian Schöneich. "A Methodology for Simultaneous Fluorogenic Derivatization and Boronate Affinity Enrichment of 3-nitrotyrosine-containing Peptides." Analytical Biochemistry 418.2 (2011): 184-96. | en_US |
dc.identifier.uri | http://hdl.handle.net/1808/22616 | |
dc.description.abstract | We synthesized and characterized a new tagging reagent, (3R,4S)-1-(4-(aminomethyl)phenylsulfonyl)pyrrolidine-3,4-diol (APPD), for the selective fluorogenic derivatization of 3-nitrotyrosine (3-NT) residues in peptides (after reduction to 3-aminotyrosine) and affinity enrichment. The synthetic 3-NT-containing peptide, FSAY(3-NO2)LER, was employed as a model for method validation. Further, this derivatization protocol was successfully tested for analysis of 3-NT-containing proteins exposed to peroxynitrite in the total protein lysate of cultured C2C12 cells. The quantitation of 3-NT content in samples was achieved through either fluorescence spectrometry or boronate affinity chromatography with detection by specific fluorescence (excitation and emission wavelengths of 360 and 510 nm, respectively); the respective limits of detection were 95 and 68 nM (19 and 13 pmol total amount) of 3-NT. Importantly, the derivatized peptides show a strong retention on a synthetic boronate affinity column, containing sulfonamide-phenylboronic acid, under mild chromatographic conditions, affording a route to separate the derivatized peptides from large amounts (milligrams) of non-derivatized peptides, and to enrich them for fluorescent detection and MS identification. Tandem MS analysis identified chemical structures of peptide 3-NT fluorescent derivatives and revealed that the fluorescent derivatives undergo efficient backbone fragmentations, permitting sequence-specific identification of protein nitration at low concentrations of 3-NT in complex protein mixtures. | en_US |
dc.publisher | Elsevier Masson | en_US |
dc.rights | This is an open access article under the terms of the Creative Commons Attribution-NonCommercial-NoDerivs License 3.0 (CC BY-NC-ND 3.0 US), which permits use and distribution in any medium, provided the original work is properly cited, the use is non-commercial and no modifications or adaptations are made. | en_US |
dc.rights.uri | http://creativecommons.org/licenses/by-nc-nd/3.0/ | |
dc.subject | 3-Nitrotyrosine | en_US |
dc.subject | (3R, 4S)-1-(4-(aminomethyl)phenylsulfonyl) pyrrolidine-3,4-diol (APPD) | en_US |
dc.subject | Fluorogenic derivatization | en_US |
dc.subject | Fluorescence | en_US |
dc.subject | Boronate affinity chromatography | en_US |
dc.subject | Mass spectrometry | en_US |
dc.subject | Proteomics | en_US |
dc.title | A Methodology for Simultaneous Fluorogenic Derivatization and Boronate Affinity Enrichment of 3-Nitrotyrosine Containing Peptides | en_US |
dc.type | Article | en_US |
kusw.kuauthor | Galeva, Nadezhda A. | |
kusw.kudepartment | Mass Spectrometry Lab | en_US |
dc.identifier.doi | 10.1016/j.ab.2011.07.024 | en_US |
kusw.oaversion | Scholarly/refereed, author accepted manuscript | en_US |
kusw.oapolicy | This item meets KU Open Access policy criteria. | en_US |
dc.rights.accessrights | openAccess |
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Except where otherwise noted, this item's license is described as: This is an open access article under the terms of the Creative Commons Attribution-NonCommercial-NoDerivs License 3.0 (CC BY-NC-ND 3.0 US), which permits use and distribution in any medium, provided the original work is properly cited, the use is non-commercial and no modifications or adaptations are made.