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Desolvation and Development of Specific Hydrophobic Core Packing during Im7 Folding

Development of a tightly packed hydrophobic core drives the folding of water-soluble globular proteins and is a key determinant of protein stability. Despite this, there remains much to be learnt about how and when the hydrophobic core becomes desolvated and tightly packed during protein folding. We...

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Autores principales: Bartlett, Alice I., Radford, Sheena E.
Formato: Texto
Lenguaje:English
Publicado: Elsevier 2010
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC2833379/
https://www.ncbi.nlm.nih.gov/pubmed/20053361
http://dx.doi.org/10.1016/j.jmb.2009.12.048
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author Bartlett, Alice I.
Radford, Sheena E.
author_facet Bartlett, Alice I.
Radford, Sheena E.
author_sort Bartlett, Alice I.
collection PubMed
description Development of a tightly packed hydrophobic core drives the folding of water-soluble globular proteins and is a key determinant of protein stability. Despite this, there remains much to be learnt about how and when the hydrophobic core becomes desolvated and tightly packed during protein folding. We have used the bacterial immunity protein Im7 to examine the specificity of hydrophobic core packing during folding. This small, four-helix protein has previously been shown to fold via a compact three-helical intermediate state. Here, overpacking substitutions, in which residue side-chain size is increased, were used to examine the specificity and malleability of core packing in the folding intermediate and rate-limiting transition state. In parallel, polar groups were introduced into the Im7 hydrophobic core via Val→Thr or Phe→Tyr substitutions and used to determine the solvation status of core residues at different stages of folding. Over 30 Im7 variants were created allowing both series of substitutions to cover all regions of the protein structure. Φ-value analysis demonstrated that the major changes in Im7 core solvation occur prior to the population of the folding intermediate, with key regions involved in docking of the short helix III remaining solvent-exposed until after the rate-limiting transition state has been traversed. In contrast, overpacking core residues revealed that some regions of the native Im7 core are remarkably malleable to increases in side-chain volume. Overpacking residues in other regions of the Im7 core result in substantial (> 2.5 kJ mol(− 1)) destabilisation of the native structure or even prevents efficient folding to the native state. This study provides new insights into Im7 folding; demonstrating that whilst desolvation occurs early during folding, adoption of a specifically packed core is achieved only at the very last step in the folding mechanism.
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spelling pubmed-28333792010-03-29 Desolvation and Development of Specific Hydrophobic Core Packing during Im7 Folding Bartlett, Alice I. Radford, Sheena E. J Mol Biol Article Development of a tightly packed hydrophobic core drives the folding of water-soluble globular proteins and is a key determinant of protein stability. Despite this, there remains much to be learnt about how and when the hydrophobic core becomes desolvated and tightly packed during protein folding. We have used the bacterial immunity protein Im7 to examine the specificity of hydrophobic core packing during folding. This small, four-helix protein has previously been shown to fold via a compact three-helical intermediate state. Here, overpacking substitutions, in which residue side-chain size is increased, were used to examine the specificity and malleability of core packing in the folding intermediate and rate-limiting transition state. In parallel, polar groups were introduced into the Im7 hydrophobic core via Val→Thr or Phe→Tyr substitutions and used to determine the solvation status of core residues at different stages of folding. Over 30 Im7 variants were created allowing both series of substitutions to cover all regions of the protein structure. Φ-value analysis demonstrated that the major changes in Im7 core solvation occur prior to the population of the folding intermediate, with key regions involved in docking of the short helix III remaining solvent-exposed until after the rate-limiting transition state has been traversed. In contrast, overpacking core residues revealed that some regions of the native Im7 core are remarkably malleable to increases in side-chain volume. Overpacking residues in other regions of the Im7 core result in substantial (> 2.5 kJ mol(− 1)) destabilisation of the native structure or even prevents efficient folding to the native state. This study provides new insights into Im7 folding; demonstrating that whilst desolvation occurs early during folding, adoption of a specifically packed core is achieved only at the very last step in the folding mechanism. Elsevier 2010-03-12 /pmc/articles/PMC2833379/ /pubmed/20053361 http://dx.doi.org/10.1016/j.jmb.2009.12.048 Text en © 2010 Elsevier Ltd. https://creativecommons.org/licenses/by/3.0/ Open Access under CC BY 3.0 (https://creativecommons.org/licenses/by/3.0/) license
spellingShingle Article
Bartlett, Alice I.
Radford, Sheena E.
Desolvation and Development of Specific Hydrophobic Core Packing during Im7 Folding
title Desolvation and Development of Specific Hydrophobic Core Packing during Im7 Folding
title_full Desolvation and Development of Specific Hydrophobic Core Packing during Im7 Folding
title_fullStr Desolvation and Development of Specific Hydrophobic Core Packing during Im7 Folding
title_full_unstemmed Desolvation and Development of Specific Hydrophobic Core Packing during Im7 Folding
title_short Desolvation and Development of Specific Hydrophobic Core Packing during Im7 Folding
title_sort desolvation and development of specific hydrophobic core packing during im7 folding
topic Article
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC2833379/
https://www.ncbi.nlm.nih.gov/pubmed/20053361
http://dx.doi.org/10.1016/j.jmb.2009.12.048
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