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A rapid and sensitive high-throughput screening method to identify compounds targeting protein–nucleic acids interactions

DNA-binding and RNA-binding proteins are usually considered ‘undruggable’ partly due to the lack of an efficient method to identify inhibitors from existing small molecule repositories. Here we report a rapid and sensitive high-throughput screening approach to identify compounds targeting protein–nu...

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Detalles Bibliográficos
Autores principales: Alonso, Nicole, Guillen, Roboan, Chambers, Jeremy W., Leng, Fenfei
Formato: Online Artículo Texto
Lenguaje:English
Publicado: Oxford University Press 2015
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4417145/
https://www.ncbi.nlm.nih.gov/pubmed/25653160
http://dx.doi.org/10.1093/nar/gkv069
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author Alonso, Nicole
Guillen, Roboan
Chambers, Jeremy W.
Leng, Fenfei
author_facet Alonso, Nicole
Guillen, Roboan
Chambers, Jeremy W.
Leng, Fenfei
author_sort Alonso, Nicole
collection PubMed
description DNA-binding and RNA-binding proteins are usually considered ‘undruggable’ partly due to the lack of an efficient method to identify inhibitors from existing small molecule repositories. Here we report a rapid and sensitive high-throughput screening approach to identify compounds targeting protein–nucleic acids interactions based on protein–DNA or protein–RNA interaction enzyme-linked immunosorbent assays (PDI-ELISA or PRI-ELISA). We validated the PDI-ELISA method using the mammalian high-mobility-group protein AT-hook 2 (HMGA2) as the protein of interest and netropsin as the inhibitor of HMGA2–DNA interactions. With this method we successfully identified several inhibitors and an activator for HMGA2–DNA interactions from a collection of 29 DNA-binding compounds. Guided by this screening excise, we showed that netropsin, the specific inhibitor of HMGA2–DNA interactions, strongly inhibited the differentiation of the mouse pre-adipocyte 3T3-L1 cells into adipocytes, most likely through a mechanism by which the inhibition is through preventing the binding of HMGA2 to the target DNA sequences. This method should be broadly applicable to identify compounds or proteins modulating many DNA-binding or RNA-binding proteins.
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spelling pubmed-44171452015-05-12 A rapid and sensitive high-throughput screening method to identify compounds targeting protein–nucleic acids interactions Alonso, Nicole Guillen, Roboan Chambers, Jeremy W. Leng, Fenfei Nucleic Acids Res Methods Online DNA-binding and RNA-binding proteins are usually considered ‘undruggable’ partly due to the lack of an efficient method to identify inhibitors from existing small molecule repositories. Here we report a rapid and sensitive high-throughput screening approach to identify compounds targeting protein–nucleic acids interactions based on protein–DNA or protein–RNA interaction enzyme-linked immunosorbent assays (PDI-ELISA or PRI-ELISA). We validated the PDI-ELISA method using the mammalian high-mobility-group protein AT-hook 2 (HMGA2) as the protein of interest and netropsin as the inhibitor of HMGA2–DNA interactions. With this method we successfully identified several inhibitors and an activator for HMGA2–DNA interactions from a collection of 29 DNA-binding compounds. Guided by this screening excise, we showed that netropsin, the specific inhibitor of HMGA2–DNA interactions, strongly inhibited the differentiation of the mouse pre-adipocyte 3T3-L1 cells into adipocytes, most likely through a mechanism by which the inhibition is through preventing the binding of HMGA2 to the target DNA sequences. This method should be broadly applicable to identify compounds or proteins modulating many DNA-binding or RNA-binding proteins. Oxford University Press 2015-04-30 2015-02-04 /pmc/articles/PMC4417145/ /pubmed/25653160 http://dx.doi.org/10.1093/nar/gkv069 Text en © The Author(s) 2015. Published by Oxford University Press on behalf of Nucleic Acids Research. http://creativecommons.org/licenses/by-nc/4.0/ This is an Open Access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by-nc/4.0/), which permits non-commercial re-use, distribution, and reproduction in any medium, provided the original work is properly cited. For commercial re-use, please contact journals.permissions@oup.com
spellingShingle Methods Online
Alonso, Nicole
Guillen, Roboan
Chambers, Jeremy W.
Leng, Fenfei
A rapid and sensitive high-throughput screening method to identify compounds targeting protein–nucleic acids interactions
title A rapid and sensitive high-throughput screening method to identify compounds targeting protein–nucleic acids interactions
title_full A rapid and sensitive high-throughput screening method to identify compounds targeting protein–nucleic acids interactions
title_fullStr A rapid and sensitive high-throughput screening method to identify compounds targeting protein–nucleic acids interactions
title_full_unstemmed A rapid and sensitive high-throughput screening method to identify compounds targeting protein–nucleic acids interactions
title_short A rapid and sensitive high-throughput screening method to identify compounds targeting protein–nucleic acids interactions
title_sort rapid and sensitive high-throughput screening method to identify compounds targeting protein–nucleic acids interactions
topic Methods Online
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4417145/
https://www.ncbi.nlm.nih.gov/pubmed/25653160
http://dx.doi.org/10.1093/nar/gkv069
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