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Characterization of early and late transition states of the folding pathway of a SH2 domain
Albeit SH2 domains are abundant protein–protein interaction modules with fundamental roles in the regulation of several physiological and molecular pathways in the cell, the available information about the determinants of their thermodynamic stability and folding properties are still very limited. I...
Autores principales: | , , , , , , , |
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Formato: | Online Artículo Texto |
Lenguaje: | English |
Publicado: |
John Wiley & Sons, Inc.
2022
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Materias: | |
Acceso en línea: | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC9112803/ https://www.ncbi.nlm.nih.gov/pubmed/35634781 http://dx.doi.org/10.1002/pro.4332 |
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author | Toto, Angelo Malagrinò, Francesca Nardella, Caterina Pennacchietti, Valeria Pagano, Livia Santorelli, Daniele Diop, Awa Gianni, Stefano |
author_facet | Toto, Angelo Malagrinò, Francesca Nardella, Caterina Pennacchietti, Valeria Pagano, Livia Santorelli, Daniele Diop, Awa Gianni, Stefano |
author_sort | Toto, Angelo |
collection | PubMed |
description | Albeit SH2 domains are abundant protein–protein interaction modules with fundamental roles in the regulation of several physiological and molecular pathways in the cell, the available information about the determinants of their thermodynamic stability and folding properties are still very limited. In this work, we provide a quantitative characterization of the folding pathway of the C‐terminal SH2 domain of SHP2, conducted through a combination of site‐directed mutagenesis and kinetic (un)folding experiments (Φ‐value analysis). The energetic profile of the folding reaction of the C‐SH2 domain is described by a three‐state mechanism characterized by the presence of two transition states and a high‐energy intermediate. The production of 29 site‐directed variants allowed us to calculate the degree of native‐like interactions occurring in the early and late events of the folding reaction. Data analysis highlights the presence of a hydrophobic folding nucleus surrounded by a lower degree of structure in the early events of folding, further consolidated as the reaction proceeds towards the native state. Interestingly, residues physically located in the functional region of the domain reported unusual Φ‐values, a hallmark of the presence of transient misfolding. We compared our results with previous ones obtained for the N‐terminal SH2 domain of SHP2. Notably, a conserved complex folding mechanism implying the presence of a folding intermediate arise from comparison, and the relative stability of such intermediate appears to be highly sequence dependent. Data are discussed under the light of previous works on SH2 domains. |
format | Online Article Text |
id | pubmed-9112803 |
institution | National Center for Biotechnology Information |
language | English |
publishDate | 2022 |
publisher | John Wiley & Sons, Inc. |
record_format | MEDLINE/PubMed |
spelling | pubmed-91128032022-05-20 Characterization of early and late transition states of the folding pathway of a SH2 domain Toto, Angelo Malagrinò, Francesca Nardella, Caterina Pennacchietti, Valeria Pagano, Livia Santorelli, Daniele Diop, Awa Gianni, Stefano Protein Sci Full‐length Papers Albeit SH2 domains are abundant protein–protein interaction modules with fundamental roles in the regulation of several physiological and molecular pathways in the cell, the available information about the determinants of their thermodynamic stability and folding properties are still very limited. In this work, we provide a quantitative characterization of the folding pathway of the C‐terminal SH2 domain of SHP2, conducted through a combination of site‐directed mutagenesis and kinetic (un)folding experiments (Φ‐value analysis). The energetic profile of the folding reaction of the C‐SH2 domain is described by a three‐state mechanism characterized by the presence of two transition states and a high‐energy intermediate. The production of 29 site‐directed variants allowed us to calculate the degree of native‐like interactions occurring in the early and late events of the folding reaction. Data analysis highlights the presence of a hydrophobic folding nucleus surrounded by a lower degree of structure in the early events of folding, further consolidated as the reaction proceeds towards the native state. Interestingly, residues physically located in the functional region of the domain reported unusual Φ‐values, a hallmark of the presence of transient misfolding. We compared our results with previous ones obtained for the N‐terminal SH2 domain of SHP2. Notably, a conserved complex folding mechanism implying the presence of a folding intermediate arise from comparison, and the relative stability of such intermediate appears to be highly sequence dependent. Data are discussed under the light of previous works on SH2 domains. John Wiley & Sons, Inc. 2022-05-17 2022-06 /pmc/articles/PMC9112803/ /pubmed/35634781 http://dx.doi.org/10.1002/pro.4332 Text en © 2022 The Authors. Protein Science published by Wiley Periodicals LLC on behalf of The Protein Society. https://creativecommons.org/licenses/by/4.0/This is an open access article under the terms of the http://creativecommons.org/licenses/by/4.0/ (https://creativecommons.org/licenses/by/4.0/) License, which permits use, distribution and reproduction in any medium, provided the original work is properly cited. |
spellingShingle | Full‐length Papers Toto, Angelo Malagrinò, Francesca Nardella, Caterina Pennacchietti, Valeria Pagano, Livia Santorelli, Daniele Diop, Awa Gianni, Stefano Characterization of early and late transition states of the folding pathway of a SH2 domain |
title | Characterization of early and late transition states of the folding pathway of a SH2 domain |
title_full | Characterization of early and late transition states of the folding pathway of a SH2 domain |
title_fullStr | Characterization of early and late transition states of the folding pathway of a SH2 domain |
title_full_unstemmed | Characterization of early and late transition states of the folding pathway of a SH2 domain |
title_short | Characterization of early and late transition states of the folding pathway of a SH2 domain |
title_sort | characterization of early and late transition states of the folding pathway of a sh2 domain |
topic | Full‐length Papers |
url | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC9112803/ https://www.ncbi.nlm.nih.gov/pubmed/35634781 http://dx.doi.org/10.1002/pro.4332 |
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