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The unfolding of iRFP713 in a crowded milieu

The exploring of biological processes in vitro under conditions of macromolecular crowding is a way to achieve an understanding of how these processes occur in vivo. In this work, we study the unfolding of the fluorescent probe iRFP713 in crowded environment in vitro. Previously, we showed that the...

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Autores principales: Stepanenko, Olesya V., Stepanenko, Olga V., Kuznetsova, Irina M., Turoverov, Konstantin K.
Formato: Online Artículo Texto
Lenguaje:English
Publicado: PeerJ Inc. 2019
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC6459179/
https://www.ncbi.nlm.nih.gov/pubmed/30993043
http://dx.doi.org/10.7717/peerj.6707
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author Stepanenko, Olesya V.
Stepanenko, Olga V.
Kuznetsova, Irina M.
Turoverov, Konstantin K.
author_facet Stepanenko, Olesya V.
Stepanenko, Olga V.
Kuznetsova, Irina M.
Turoverov, Konstantin K.
author_sort Stepanenko, Olesya V.
collection PubMed
description The exploring of biological processes in vitro under conditions of macromolecular crowding is a way to achieve an understanding of how these processes occur in vivo. In this work, we study the unfolding of the fluorescent probe iRFP713 in crowded environment in vitro. Previously, we showed that the unfolding of the dimeric iRFP713 is accompanied by the formation of a compact monomer and an intermediate state of the protein. In the intermediate state, the macromolecules of iRFP713 have hydrophobic clusters exposed to the surface of the protein and are prone to aggregation. Concentrated solutions of polyethylene glycol (PEG-8000), Dextran-40 and Dextran-70 with a molecular mass of 8000, 40000 and 70000 Da, respectively, were used to model the conditions for macromolecular crowding. A limited available space provided by all the crowding agents used favors to the enhanced aggregation of iRFP713 in the intermediate state at the concentration of guanidine hydrochloride (GdnHCl), at which the charge of protein surface is neutralized by the guanidine cations. This is in line with the theory of the excluded volume. In concentrated solutions of the crowding agents (240–300 mg/ml), the stabilization of the structure of iRFP713 in the intermediate state is observed. PEG-8000 also enhances the stability of iRFP713 in the monomeric compact state, whereas in concentrated solutions of Dextran-40 and Dextran-70 the resistance of the protein in the monomeric state against GdnHCl-induced unfolding decreases. The obtained data argues for the excluded volume effect being not the only factor that contributes the behavior of biological molecules in a crowded milieu. Crowding agents do not affect the structure of the native dimer of iRFP713, which excludes the direct interactions between the target protein and the crowding agents. PEGs of different molecular mass and Dextran-40/Dextran-70 are known to influence the solvent properties of water. The solvent dipolarity/polarizability and basicity/acidity in aqueous solutions of these crowding agents vary in different ways. The change of the solvent properties in aqueous solutions of crowding agents might impact the functioning of a target protein.
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spelling pubmed-64591792019-04-16 The unfolding of iRFP713 in a crowded milieu Stepanenko, Olesya V. Stepanenko, Olga V. Kuznetsova, Irina M. Turoverov, Konstantin K. PeerJ Biochemistry The exploring of biological processes in vitro under conditions of macromolecular crowding is a way to achieve an understanding of how these processes occur in vivo. In this work, we study the unfolding of the fluorescent probe iRFP713 in crowded environment in vitro. Previously, we showed that the unfolding of the dimeric iRFP713 is accompanied by the formation of a compact monomer and an intermediate state of the protein. In the intermediate state, the macromolecules of iRFP713 have hydrophobic clusters exposed to the surface of the protein and are prone to aggregation. Concentrated solutions of polyethylene glycol (PEG-8000), Dextran-40 and Dextran-70 with a molecular mass of 8000, 40000 and 70000 Da, respectively, were used to model the conditions for macromolecular crowding. A limited available space provided by all the crowding agents used favors to the enhanced aggregation of iRFP713 in the intermediate state at the concentration of guanidine hydrochloride (GdnHCl), at which the charge of protein surface is neutralized by the guanidine cations. This is in line with the theory of the excluded volume. In concentrated solutions of the crowding agents (240–300 mg/ml), the stabilization of the structure of iRFP713 in the intermediate state is observed. PEG-8000 also enhances the stability of iRFP713 in the monomeric compact state, whereas in concentrated solutions of Dextran-40 and Dextran-70 the resistance of the protein in the monomeric state against GdnHCl-induced unfolding decreases. The obtained data argues for the excluded volume effect being not the only factor that contributes the behavior of biological molecules in a crowded milieu. Crowding agents do not affect the structure of the native dimer of iRFP713, which excludes the direct interactions between the target protein and the crowding agents. PEGs of different molecular mass and Dextran-40/Dextran-70 are known to influence the solvent properties of water. The solvent dipolarity/polarizability and basicity/acidity in aqueous solutions of these crowding agents vary in different ways. The change of the solvent properties in aqueous solutions of crowding agents might impact the functioning of a target protein. PeerJ Inc. 2019-04-08 /pmc/articles/PMC6459179/ /pubmed/30993043 http://dx.doi.org/10.7717/peerj.6707 Text en ©2019 Stepanenko et al. http://creativecommons.org/licenses/by/4.0/ This is an open access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/4.0/) , which permits unrestricted use, distribution, reproduction and adaptation in any medium and for any purpose provided that it is properly attributed. For attribution, the original author(s), title, publication source (PeerJ) and either DOI or URL of the article must be cited.
spellingShingle Biochemistry
Stepanenko, Olesya V.
Stepanenko, Olga V.
Kuznetsova, Irina M.
Turoverov, Konstantin K.
The unfolding of iRFP713 in a crowded milieu
title The unfolding of iRFP713 in a crowded milieu
title_full The unfolding of iRFP713 in a crowded milieu
title_fullStr The unfolding of iRFP713 in a crowded milieu
title_full_unstemmed The unfolding of iRFP713 in a crowded milieu
title_short The unfolding of iRFP713 in a crowded milieu
title_sort unfolding of irfp713 in a crowded milieu
topic Biochemistry
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC6459179/
https://www.ncbi.nlm.nih.gov/pubmed/30993043
http://dx.doi.org/10.7717/peerj.6707
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