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Protein Binder (ProBi) as a New Class of Structurally Robust Non-Antibody Protein Scaffold for Directed Evolution

Engineered small non-antibody protein scaffolds are a promising alternative to antibodies and are especially attractive for use in protein therapeutics and diagnostics. The advantages include smaller size and a more robust, single-domain structural framework with a defined binding surface amenable t...

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Autores principales: Pham, Phuong Ngoc, Huličiak, Maroš, Biedermannová, Lada, Černý, Jiří, Charnavets, Tatsiana, Fuertes, Gustavo, Herynek, Štěpán, Kolářová, Lucie, Kolenko, Petr, Pavlíček, Jiří, Zahradník, Jiří, Mikulecky, Pavel, Schneider, Bohdan
Formato: Online Artículo Texto
Lenguaje:English
Publicado: MDPI 2021
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC7911045/
https://www.ncbi.nlm.nih.gov/pubmed/33514045
http://dx.doi.org/10.3390/v13020190
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author Pham, Phuong Ngoc
Huličiak, Maroš
Biedermannová, Lada
Černý, Jiří
Charnavets, Tatsiana
Fuertes, Gustavo
Herynek, Štěpán
Kolářová, Lucie
Kolenko, Petr
Pavlíček, Jiří
Zahradník, Jiří
Mikulecky, Pavel
Schneider, Bohdan
author_facet Pham, Phuong Ngoc
Huličiak, Maroš
Biedermannová, Lada
Černý, Jiří
Charnavets, Tatsiana
Fuertes, Gustavo
Herynek, Štěpán
Kolářová, Lucie
Kolenko, Petr
Pavlíček, Jiří
Zahradník, Jiří
Mikulecky, Pavel
Schneider, Bohdan
author_sort Pham, Phuong Ngoc
collection PubMed
description Engineered small non-antibody protein scaffolds are a promising alternative to antibodies and are especially attractive for use in protein therapeutics and diagnostics. The advantages include smaller size and a more robust, single-domain structural framework with a defined binding surface amenable to mutation. This calls for a more systematic approach in designing new scaffolds suitable for use in one or more methods of directed evolution. We hereby describe a process based on an analysis of protein structures from the Protein Data Bank and their experimental examination. The candidate protein scaffolds were subjected to a thorough screening including computational evaluation of the mutability, and experimental determination of their expression yield in E. coli, solubility, and thermostability. In the next step, we examined several variants of the candidate scaffolds including their wild types and alanine mutants. We proved the applicability of this systematic procedure by selecting a monomeric single-domain human protein with a fold different from previously known scaffolds. The newly developed scaffold, called ProBi (Protein Binder), contains two independently mutable surface patches. We demonstrated its functionality by training it as a binder against human interleukin-10, a medically important cytokine. The procedure yielded scaffold-related variants with nanomolar affinity.
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spelling pubmed-79110452021-02-28 Protein Binder (ProBi) as a New Class of Structurally Robust Non-Antibody Protein Scaffold for Directed Evolution Pham, Phuong Ngoc Huličiak, Maroš Biedermannová, Lada Černý, Jiří Charnavets, Tatsiana Fuertes, Gustavo Herynek, Štěpán Kolářová, Lucie Kolenko, Petr Pavlíček, Jiří Zahradník, Jiří Mikulecky, Pavel Schneider, Bohdan Viruses Article Engineered small non-antibody protein scaffolds are a promising alternative to antibodies and are especially attractive for use in protein therapeutics and diagnostics. The advantages include smaller size and a more robust, single-domain structural framework with a defined binding surface amenable to mutation. This calls for a more systematic approach in designing new scaffolds suitable for use in one or more methods of directed evolution. We hereby describe a process based on an analysis of protein structures from the Protein Data Bank and their experimental examination. The candidate protein scaffolds were subjected to a thorough screening including computational evaluation of the mutability, and experimental determination of their expression yield in E. coli, solubility, and thermostability. In the next step, we examined several variants of the candidate scaffolds including their wild types and alanine mutants. We proved the applicability of this systematic procedure by selecting a monomeric single-domain human protein with a fold different from previously known scaffolds. The newly developed scaffold, called ProBi (Protein Binder), contains two independently mutable surface patches. We demonstrated its functionality by training it as a binder against human interleukin-10, a medically important cytokine. The procedure yielded scaffold-related variants with nanomolar affinity. MDPI 2021-01-27 /pmc/articles/PMC7911045/ /pubmed/33514045 http://dx.doi.org/10.3390/v13020190 Text en © 2021 by the authors. https://creativecommons.org/licenses/by/4.0/Licensee MDPI, Basel, Switzerland. This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY) license (http://creativecommons.org/licenses/by/4.0/ (https://creativecommons.org/licenses/by/4.0/) ).
spellingShingle Article
Pham, Phuong Ngoc
Huličiak, Maroš
Biedermannová, Lada
Černý, Jiří
Charnavets, Tatsiana
Fuertes, Gustavo
Herynek, Štěpán
Kolářová, Lucie
Kolenko, Petr
Pavlíček, Jiří
Zahradník, Jiří
Mikulecky, Pavel
Schneider, Bohdan
Protein Binder (ProBi) as a New Class of Structurally Robust Non-Antibody Protein Scaffold for Directed Evolution
title Protein Binder (ProBi) as a New Class of Structurally Robust Non-Antibody Protein Scaffold for Directed Evolution
title_full Protein Binder (ProBi) as a New Class of Structurally Robust Non-Antibody Protein Scaffold for Directed Evolution
title_fullStr Protein Binder (ProBi) as a New Class of Structurally Robust Non-Antibody Protein Scaffold for Directed Evolution
title_full_unstemmed Protein Binder (ProBi) as a New Class of Structurally Robust Non-Antibody Protein Scaffold for Directed Evolution
title_short Protein Binder (ProBi) as a New Class of Structurally Robust Non-Antibody Protein Scaffold for Directed Evolution
title_sort protein binder (probi) as a new class of structurally robust non-antibody protein scaffold for directed evolution
topic Article
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC7911045/
https://www.ncbi.nlm.nih.gov/pubmed/33514045
http://dx.doi.org/10.3390/v13020190
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