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Toward a Rationale for the PTC124 (Ataluren) Promoted Readthrough of Premature Stop Codons: A Computational Approach and GFP-Reporter Cell-Based Assay
[Image: see text] The presence in the mRNA of premature stop codons (PTCs) results in protein truncation responsible for several inherited (genetic) diseases. A well-known example of these diseases is cystic fibrosis (CF), where approximately 10% (worldwide) of patients have nonsense mutations in th...
Autores principales: | , , , , , , , |
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Formato: | Online Artículo Texto |
Lenguaje: | English |
Publicado: |
American
Chemical
Society
2014
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Acceso en línea: | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4167060/ https://www.ncbi.nlm.nih.gov/pubmed/24483936 http://dx.doi.org/10.1021/mp400230s |
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author | Lentini, Laura Melfi, Raffaella Di Leonardo, Aldo Spinello, Angelo Barone, Giampaolo Pace, Andrea Palumbo Piccionello, Antonio Pibiri, Ivana |
author_facet | Lentini, Laura Melfi, Raffaella Di Leonardo, Aldo Spinello, Angelo Barone, Giampaolo Pace, Andrea Palumbo Piccionello, Antonio Pibiri, Ivana |
author_sort | Lentini, Laura |
collection | PubMed |
description | [Image: see text] The presence in the mRNA of premature stop codons (PTCs) results in protein truncation responsible for several inherited (genetic) diseases. A well-known example of these diseases is cystic fibrosis (CF), where approximately 10% (worldwide) of patients have nonsense mutations in the CF transmembrane regulator (CFTR) gene. PTC124 (3-(5-(2-fluorophenyl)-1,2,4-oxadiazol-3-yl)-benzoic acid), also known as Ataluren, is a small molecule that has been suggested to allow PTC readthrough even though its target has yet to be identified. In the lack of a general consensus about its mechanism of action, we experimentally tested the ability of PTC124 to promote the readthrough of premature termination codons by using a new reporter. The reporter vector was based on a plasmid harboring the H2B histone coding sequence fused in frame with the green fluorescent protein (GFP) cDNA, and a TGA stop codon was introduced in the H2B-GFP gene by site-directed mutagenesis. Additionally, an unprecedented computational study on the putative supramolecular interaction between PTC124 and an 11-codon (33-nucleotides) sequence corresponding to a CFTR mRNA fragment containing a central UGA nonsense mutation showed a specific interaction between PTC124 and the UGA codon. Altogether, the H2B-GFP-opal based assay and the molecular dynamics (MD) simulation support the hypothesis that PTC124 is able to promote the specific readthrough of internal TGA premature stop codons. |
format | Online Article Text |
id | pubmed-4167060 |
institution | National Center for Biotechnology Information |
language | English |
publishDate | 2014 |
publisher | American
Chemical
Society |
record_format | MEDLINE/PubMed |
spelling | pubmed-41670602015-01-31 Toward a Rationale for the PTC124 (Ataluren) Promoted Readthrough of Premature Stop Codons: A Computational Approach and GFP-Reporter Cell-Based Assay Lentini, Laura Melfi, Raffaella Di Leonardo, Aldo Spinello, Angelo Barone, Giampaolo Pace, Andrea Palumbo Piccionello, Antonio Pibiri, Ivana Mol Pharm [Image: see text] The presence in the mRNA of premature stop codons (PTCs) results in protein truncation responsible for several inherited (genetic) diseases. A well-known example of these diseases is cystic fibrosis (CF), where approximately 10% (worldwide) of patients have nonsense mutations in the CF transmembrane regulator (CFTR) gene. PTC124 (3-(5-(2-fluorophenyl)-1,2,4-oxadiazol-3-yl)-benzoic acid), also known as Ataluren, is a small molecule that has been suggested to allow PTC readthrough even though its target has yet to be identified. In the lack of a general consensus about its mechanism of action, we experimentally tested the ability of PTC124 to promote the readthrough of premature termination codons by using a new reporter. The reporter vector was based on a plasmid harboring the H2B histone coding sequence fused in frame with the green fluorescent protein (GFP) cDNA, and a TGA stop codon was introduced in the H2B-GFP gene by site-directed mutagenesis. Additionally, an unprecedented computational study on the putative supramolecular interaction between PTC124 and an 11-codon (33-nucleotides) sequence corresponding to a CFTR mRNA fragment containing a central UGA nonsense mutation showed a specific interaction between PTC124 and the UGA codon. Altogether, the H2B-GFP-opal based assay and the molecular dynamics (MD) simulation support the hypothesis that PTC124 is able to promote the specific readthrough of internal TGA premature stop codons. American Chemical Society 2014-01-31 2014-03-03 /pmc/articles/PMC4167060/ /pubmed/24483936 http://dx.doi.org/10.1021/mp400230s Text en Copyright © 2014 American Chemical Society Terms of Use (http://pubs.acs.org/page/policy/authorchoice_termsofuse.html) |
spellingShingle | Lentini, Laura Melfi, Raffaella Di Leonardo, Aldo Spinello, Angelo Barone, Giampaolo Pace, Andrea Palumbo Piccionello, Antonio Pibiri, Ivana Toward a Rationale for the PTC124 (Ataluren) Promoted Readthrough of Premature Stop Codons: A Computational Approach and GFP-Reporter Cell-Based Assay |
title | Toward a Rationale for the PTC124 (Ataluren) Promoted
Readthrough of Premature Stop Codons: A Computational Approach and
GFP-Reporter Cell-Based Assay |
title_full | Toward a Rationale for the PTC124 (Ataluren) Promoted
Readthrough of Premature Stop Codons: A Computational Approach and
GFP-Reporter Cell-Based Assay |
title_fullStr | Toward a Rationale for the PTC124 (Ataluren) Promoted
Readthrough of Premature Stop Codons: A Computational Approach and
GFP-Reporter Cell-Based Assay |
title_full_unstemmed | Toward a Rationale for the PTC124 (Ataluren) Promoted
Readthrough of Premature Stop Codons: A Computational Approach and
GFP-Reporter Cell-Based Assay |
title_short | Toward a Rationale for the PTC124 (Ataluren) Promoted
Readthrough of Premature Stop Codons: A Computational Approach and
GFP-Reporter Cell-Based Assay |
title_sort | toward a rationale for the ptc124 (ataluren) promoted
readthrough of premature stop codons: a computational approach and
gfp-reporter cell-based assay |
url | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4167060/ https://www.ncbi.nlm.nih.gov/pubmed/24483936 http://dx.doi.org/10.1021/mp400230s |
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