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A new soaking procedure for X-ray crystallographic structural determination of protein–peptide complexes

Interactions between a protein and a peptide motif of its protein partner are prevalent in nature. Often, a protein also has multiple interaction partners. X-ray protein crystallography is commonly used to examine these interactions in terms of bond distances and angles as well as to describe hotspo...

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Autores principales: Ballone, Alice, Lau, Roxanne A., Zweipfenning, Fabian P. A., Ottmann, Christian
Formato: Online Artículo Texto
Lenguaje:English
Publicado: International Union of Crystallography 2020
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC7531243/
https://www.ncbi.nlm.nih.gov/pubmed/33006579
http://dx.doi.org/10.1107/S2053230X2001122X
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author Ballone, Alice
Lau, Roxanne A.
Zweipfenning, Fabian P. A.
Ottmann, Christian
author_facet Ballone, Alice
Lau, Roxanne A.
Zweipfenning, Fabian P. A.
Ottmann, Christian
author_sort Ballone, Alice
collection PubMed
description Interactions between a protein and a peptide motif of its protein partner are prevalent in nature. Often, a protein also has multiple interaction partners. X-ray protein crystallography is commonly used to examine these interactions in terms of bond distances and angles as well as to describe hotspots within protein complexes. However, the crystallization process presents a significant bottleneck in structure determination since it often requires notably time-consuming screening procedures, which involve testing a broad range of crystallization conditions via a trial-and-error approach. This difficulty is also increased as each protein–peptide complex does not necessarily crystallize under the same conditions. Here, a new co-crystallization/peptide-soaking method is presented which circumvents the need to return to the initial lengthy crystal screening and optimization processes for each consequent new complex. The 14-3-3σ protein, which has multiple interacting partners with specific peptidic motifs, was used as a case study. It was found that co-crystals of 14-3-3σ and a low-affinity peptide from one of its partners, c-Jun, could easily be soaked with another interacting peptide to quickly and easily generate new structures at high resolution. Not only does this significantly reduce the production time, but new 14-3-3–peptide structures that were previously not accessible with the 14-3-3σ isoform, despite screening hundreds of other different conditions, were now also able to be resolved. The findings achieved in this study may be considered as a supporting and practical guide to potentially enable the acceleration of the crystallization process of any protein–peptide system.
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spelling pubmed-75312432020-10-27 A new soaking procedure for X-ray crystallographic structural determination of protein–peptide complexes Ballone, Alice Lau, Roxanne A. Zweipfenning, Fabian P. A. Ottmann, Christian Acta Crystallogr F Struct Biol Commun Method Communications Interactions between a protein and a peptide motif of its protein partner are prevalent in nature. Often, a protein also has multiple interaction partners. X-ray protein crystallography is commonly used to examine these interactions in terms of bond distances and angles as well as to describe hotspots within protein complexes. However, the crystallization process presents a significant bottleneck in structure determination since it often requires notably time-consuming screening procedures, which involve testing a broad range of crystallization conditions via a trial-and-error approach. This difficulty is also increased as each protein–peptide complex does not necessarily crystallize under the same conditions. Here, a new co-crystallization/peptide-soaking method is presented which circumvents the need to return to the initial lengthy crystal screening and optimization processes for each consequent new complex. The 14-3-3σ protein, which has multiple interacting partners with specific peptidic motifs, was used as a case study. It was found that co-crystals of 14-3-3σ and a low-affinity peptide from one of its partners, c-Jun, could easily be soaked with another interacting peptide to quickly and easily generate new structures at high resolution. Not only does this significantly reduce the production time, but new 14-3-3–peptide structures that were previously not accessible with the 14-3-3σ isoform, despite screening hundreds of other different conditions, were now also able to be resolved. The findings achieved in this study may be considered as a supporting and practical guide to potentially enable the acceleration of the crystallization process of any protein–peptide system. International Union of Crystallography 2020-09-15 /pmc/articles/PMC7531243/ /pubmed/33006579 http://dx.doi.org/10.1107/S2053230X2001122X Text en © Ballone et al. 2020 http://creativecommons.org/licenses/by/4.0/ This is an open-access article distributed under the terms of the Creative Commons Attribution (CC-BY) Licence, which permits unrestricted use, distribution, and reproduction in any medium, provided the original authors and source are cited.http://creativecommons.org/licenses/by/4.0/
spellingShingle Method Communications
Ballone, Alice
Lau, Roxanne A.
Zweipfenning, Fabian P. A.
Ottmann, Christian
A new soaking procedure for X-ray crystallographic structural determination of protein–peptide complexes
title A new soaking procedure for X-ray crystallographic structural determination of protein–peptide complexes
title_full A new soaking procedure for X-ray crystallographic structural determination of protein–peptide complexes
title_fullStr A new soaking procedure for X-ray crystallographic structural determination of protein–peptide complexes
title_full_unstemmed A new soaking procedure for X-ray crystallographic structural determination of protein–peptide complexes
title_short A new soaking procedure for X-ray crystallographic structural determination of protein–peptide complexes
title_sort new soaking procedure for x-ray crystallographic structural determination of protein–peptide complexes
topic Method Communications
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC7531243/
https://www.ncbi.nlm.nih.gov/pubmed/33006579
http://dx.doi.org/10.1107/S2053230X2001122X
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