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Creating a Selective Nanobody Against 3-Nitrotyrosine Containing Proteins

A critical step in developing therapeutics for oxidative stress-related pathologies is the ability to determine which specific modified protein species are innocuous by-products of pathology and which are causative agents. To achieve this goal, technologies are needed that can identify, characterize...

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Autores principales: Van Fossen, Elise M., Grutzius, Sonia, Ruby, Carl E., Mourich, Dan V., Cebra, Chris, Bracha, Shay, Karplus, P. Andrew, Cooley, Richard B., Mehl, Ryan A.
Formato: Online Artículo Texto
Lenguaje:English
Publicado: Frontiers Media S.A. 2022
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC8899190/
https://www.ncbi.nlm.nih.gov/pubmed/35265586
http://dx.doi.org/10.3389/fchem.2022.835229
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author Van Fossen, Elise M.
Grutzius, Sonia
Ruby, Carl E.
Mourich, Dan V.
Cebra, Chris
Bracha, Shay
Karplus, P. Andrew
Cooley, Richard B.
Mehl, Ryan A.
author_facet Van Fossen, Elise M.
Grutzius, Sonia
Ruby, Carl E.
Mourich, Dan V.
Cebra, Chris
Bracha, Shay
Karplus, P. Andrew
Cooley, Richard B.
Mehl, Ryan A.
author_sort Van Fossen, Elise M.
collection PubMed
description A critical step in developing therapeutics for oxidative stress-related pathologies is the ability to determine which specific modified protein species are innocuous by-products of pathology and which are causative agents. To achieve this goal, technologies are needed that can identify, characterize and quantify oxidative post translational modifications (oxPTMs). Nanobodies (Nbs) represent exquisite tools for intracellular tracking of molecules due to their small size, stability and engineerability. Here, we demonstrate that it is possible to develop a selective Nb against an oxPTM protein, with the key advance being the use of genetic code expansion (GCE) to provide an efficient source of the large quantities of high-quality, homogenous and site-specific oxPTM-containing protein needed for the Nb selection process. In this proof-of-concept study, we produce a Nb selective for a 3-nitrotyrosine (nitroTyr) modified form of the 14-3-3 signaling protein with a lesser recognition of nitroTyr in other protein contexts. This advance opens the door to the GCE-facilitated development of other anti-PTM Nbs.
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spelling pubmed-88991902022-03-08 Creating a Selective Nanobody Against 3-Nitrotyrosine Containing Proteins Van Fossen, Elise M. Grutzius, Sonia Ruby, Carl E. Mourich, Dan V. Cebra, Chris Bracha, Shay Karplus, P. Andrew Cooley, Richard B. Mehl, Ryan A. Front Chem Chemistry A critical step in developing therapeutics for oxidative stress-related pathologies is the ability to determine which specific modified protein species are innocuous by-products of pathology and which are causative agents. To achieve this goal, technologies are needed that can identify, characterize and quantify oxidative post translational modifications (oxPTMs). Nanobodies (Nbs) represent exquisite tools for intracellular tracking of molecules due to their small size, stability and engineerability. Here, we demonstrate that it is possible to develop a selective Nb against an oxPTM protein, with the key advance being the use of genetic code expansion (GCE) to provide an efficient source of the large quantities of high-quality, homogenous and site-specific oxPTM-containing protein needed for the Nb selection process. In this proof-of-concept study, we produce a Nb selective for a 3-nitrotyrosine (nitroTyr) modified form of the 14-3-3 signaling protein with a lesser recognition of nitroTyr in other protein contexts. This advance opens the door to the GCE-facilitated development of other anti-PTM Nbs. Frontiers Media S.A. 2022-02-21 /pmc/articles/PMC8899190/ /pubmed/35265586 http://dx.doi.org/10.3389/fchem.2022.835229 Text en Copyright © 2022 Van Fossen, Grutzius, Ruby, Mourich, Cebra, Bracha, Karplus, Cooley and Mehl. https://creativecommons.org/licenses/by/4.0/This is an open-access article distributed under the terms of the Creative Commons Attribution License (CC BY). The use, distribution or reproduction in other forums is permitted, provided the original author(s) and the copyright owner(s) are credited and that the original publication in this journal is cited, in accordance with accepted academic practice. No use, distribution or reproduction is permitted which does not comply with these terms.
spellingShingle Chemistry
Van Fossen, Elise M.
Grutzius, Sonia
Ruby, Carl E.
Mourich, Dan V.
Cebra, Chris
Bracha, Shay
Karplus, P. Andrew
Cooley, Richard B.
Mehl, Ryan A.
Creating a Selective Nanobody Against 3-Nitrotyrosine Containing Proteins
title Creating a Selective Nanobody Against 3-Nitrotyrosine Containing Proteins
title_full Creating a Selective Nanobody Against 3-Nitrotyrosine Containing Proteins
title_fullStr Creating a Selective Nanobody Against 3-Nitrotyrosine Containing Proteins
title_full_unstemmed Creating a Selective Nanobody Against 3-Nitrotyrosine Containing Proteins
title_short Creating a Selective Nanobody Against 3-Nitrotyrosine Containing Proteins
title_sort creating a selective nanobody against 3-nitrotyrosine containing proteins
topic Chemistry
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC8899190/
https://www.ncbi.nlm.nih.gov/pubmed/35265586
http://dx.doi.org/10.3389/fchem.2022.835229
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