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Exploring the binding of rationally engineered tandem-repeat proteins to E3 ubiquitin ligase Keap1

The process of displaying functional peptides by ‘grafting’ them onto loops of a stable protein scaffold can be used to impart binding affinity for a target, but it can be difficult to predict the affinity of the grafted peptide and the effect of grafting on scaffold stability. In this study, we sho...

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Detalles Bibliográficos
Autores principales: Madden, Sarah K, Itzhaki, Laura S
Formato: Online Artículo Texto
Lenguaje:English
Publicado: Oxford University Press 2021
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC8660007/
https://www.ncbi.nlm.nih.gov/pubmed/34882773
http://dx.doi.org/10.1093/protein/gzab027
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author Madden, Sarah K
Itzhaki, Laura S
author_facet Madden, Sarah K
Itzhaki, Laura S
author_sort Madden, Sarah K
collection PubMed
description The process of displaying functional peptides by ‘grafting’ them onto loops of a stable protein scaffold can be used to impart binding affinity for a target, but it can be difficult to predict the affinity of the grafted peptide and the effect of grafting on scaffold stability. In this study, we show that a series of peptides that bind to the E3 ubiquitin ligase Keap1 can be grafted into the inter-repeat loop of a consensus-designed tetratricopeptide repeat (CTPR) protein resulting in proteins with high stability. We found that these CTPR-grafted peptides had similar affinities to their free peptide counterparts and achieved a low nanomolar range. This result is likely due to a good structural match between the inter-repeat loop of the CTPR and the Keap1-binding peptide. The grafting process led to the discovery of a new Keap1-binding peptide, Ac-LDPETGELL-NH(2,) with low nanomolar affinity for Keap1, highlighting the potential of the repeat-protein class for application in peptide display.
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spelling pubmed-86600072021-12-10 Exploring the binding of rationally engineered tandem-repeat proteins to E3 ubiquitin ligase Keap1 Madden, Sarah K Itzhaki, Laura S Protein Eng Des Sel Original Article The process of displaying functional peptides by ‘grafting’ them onto loops of a stable protein scaffold can be used to impart binding affinity for a target, but it can be difficult to predict the affinity of the grafted peptide and the effect of grafting on scaffold stability. In this study, we show that a series of peptides that bind to the E3 ubiquitin ligase Keap1 can be grafted into the inter-repeat loop of a consensus-designed tetratricopeptide repeat (CTPR) protein resulting in proteins with high stability. We found that these CTPR-grafted peptides had similar affinities to their free peptide counterparts and achieved a low nanomolar range. This result is likely due to a good structural match between the inter-repeat loop of the CTPR and the Keap1-binding peptide. The grafting process led to the discovery of a new Keap1-binding peptide, Ac-LDPETGELL-NH(2,) with low nanomolar affinity for Keap1, highlighting the potential of the repeat-protein class for application in peptide display. Oxford University Press 2021-12-09 /pmc/articles/PMC8660007/ /pubmed/34882773 http://dx.doi.org/10.1093/protein/gzab027 Text en © The Author(s) 2021. Published by Oxford University Press. https://creativecommons.org/licenses/by/4.0/This is an Open Access article distributed under the terms of the Creative Commons Attribution License (https://creativecommons.org/licenses/by/4.0/), which permits unrestricted reuse, distribution, and reproduction in any medium, provided the original work is properly cited.
spellingShingle Original Article
Madden, Sarah K
Itzhaki, Laura S
Exploring the binding of rationally engineered tandem-repeat proteins to E3 ubiquitin ligase Keap1
title Exploring the binding of rationally engineered tandem-repeat proteins to E3 ubiquitin ligase Keap1
title_full Exploring the binding of rationally engineered tandem-repeat proteins to E3 ubiquitin ligase Keap1
title_fullStr Exploring the binding of rationally engineered tandem-repeat proteins to E3 ubiquitin ligase Keap1
title_full_unstemmed Exploring the binding of rationally engineered tandem-repeat proteins to E3 ubiquitin ligase Keap1
title_short Exploring the binding of rationally engineered tandem-repeat proteins to E3 ubiquitin ligase Keap1
title_sort exploring the binding of rationally engineered tandem-repeat proteins to e3 ubiquitin ligase keap1
topic Original Article
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC8660007/
https://www.ncbi.nlm.nih.gov/pubmed/34882773
http://dx.doi.org/10.1093/protein/gzab027
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