Cargando…
Rational engineering of the Neurospora VS ribozyme to allow substrate recognition via different kissing-loop interactions
The Neurospora VS ribozyme is a catalytic RNA that has the unique ability to specifically recognize and cleave a stem-loop substrate through formation of a highly stable kissing-loop interaction (KLI). In order to explore the engineering potential of the VS ribozyme to cleave alternate substrates, w...
Autores principales: | , , , |
---|---|
Formato: | Online Artículo Texto |
Lenguaje: | English |
Publicado: |
Oxford University Press
2016
|
Materias: | |
Acceso en línea: | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5001590/ https://www.ncbi.nlm.nih.gov/pubmed/27166370 http://dx.doi.org/10.1093/nar/gkw401 |
_version_ | 1782450446020902912 |
---|---|
author | Lacroix-Labonté, Julie Girard, Nicolas Dagenais, Pierre Legault, Pascale |
author_facet | Lacroix-Labonté, Julie Girard, Nicolas Dagenais, Pierre Legault, Pascale |
author_sort | Lacroix-Labonté, Julie |
collection | PubMed |
description | The Neurospora VS ribozyme is a catalytic RNA that has the unique ability to specifically recognize and cleave a stem-loop substrate through formation of a highly stable kissing-loop interaction (KLI). In order to explore the engineering potential of the VS ribozyme to cleave alternate substrates, we substituted the wild-type KLI by other known KLIs using an innovative engineering method that combines rational and combinatorial approaches. A bioinformatic search of the protein data bank was initially performed to identify KLIs that are structurally similar to the one found in the VS ribozyme. Next, substrate/ribozyme (S/R) pairs that incorporate these alternative KLIs were kinetically and structurally characterized. Interestingly, several of the resulting S/R pairs allowed substrate cleavage with substantial catalytic efficiency, although with reduced activity compared to the reference S/R pair. Overall, this study describes an innovative approach for RNA engineering and establishes that the KLI of the trans VS ribozyme can be adapted to cleave other folded RNA substrates. |
format | Online Article Text |
id | pubmed-5001590 |
institution | National Center for Biotechnology Information |
language | English |
publishDate | 2016 |
publisher | Oxford University Press |
record_format | MEDLINE/PubMed |
spelling | pubmed-50015902016-12-07 Rational engineering of the Neurospora VS ribozyme to allow substrate recognition via different kissing-loop interactions Lacroix-Labonté, Julie Girard, Nicolas Dagenais, Pierre Legault, Pascale Nucleic Acids Res RNA The Neurospora VS ribozyme is a catalytic RNA that has the unique ability to specifically recognize and cleave a stem-loop substrate through formation of a highly stable kissing-loop interaction (KLI). In order to explore the engineering potential of the VS ribozyme to cleave alternate substrates, we substituted the wild-type KLI by other known KLIs using an innovative engineering method that combines rational and combinatorial approaches. A bioinformatic search of the protein data bank was initially performed to identify KLIs that are structurally similar to the one found in the VS ribozyme. Next, substrate/ribozyme (S/R) pairs that incorporate these alternative KLIs were kinetically and structurally characterized. Interestingly, several of the resulting S/R pairs allowed substrate cleavage with substantial catalytic efficiency, although with reduced activity compared to the reference S/R pair. Overall, this study describes an innovative approach for RNA engineering and establishes that the KLI of the trans VS ribozyme can be adapted to cleave other folded RNA substrates. Oxford University Press 2016-08-19 2016-05-10 /pmc/articles/PMC5001590/ /pubmed/27166370 http://dx.doi.org/10.1093/nar/gkw401 Text en © The Author(s) 2016. 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 | RNA Lacroix-Labonté, Julie Girard, Nicolas Dagenais, Pierre Legault, Pascale Rational engineering of the Neurospora VS ribozyme to allow substrate recognition via different kissing-loop interactions |
title | Rational engineering of the Neurospora VS ribozyme to allow substrate recognition via different kissing-loop interactions |
title_full | Rational engineering of the Neurospora VS ribozyme to allow substrate recognition via different kissing-loop interactions |
title_fullStr | Rational engineering of the Neurospora VS ribozyme to allow substrate recognition via different kissing-loop interactions |
title_full_unstemmed | Rational engineering of the Neurospora VS ribozyme to allow substrate recognition via different kissing-loop interactions |
title_short | Rational engineering of the Neurospora VS ribozyme to allow substrate recognition via different kissing-loop interactions |
title_sort | rational engineering of the neurospora vs ribozyme to allow substrate recognition via different kissing-loop interactions |
topic | RNA |
url | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5001590/ https://www.ncbi.nlm.nih.gov/pubmed/27166370 http://dx.doi.org/10.1093/nar/gkw401 |
work_keys_str_mv | AT lacroixlabontejulie rationalengineeringoftheneurosporavsribozymetoallowsubstraterecognitionviadifferentkissingloopinteractions AT girardnicolas rationalengineeringoftheneurosporavsribozymetoallowsubstraterecognitionviadifferentkissingloopinteractions AT dagenaispierre rationalengineeringoftheneurosporavsribozymetoallowsubstraterecognitionviadifferentkissingloopinteractions AT legaultpascale rationalengineeringoftheneurosporavsribozymetoallowsubstraterecognitionviadifferentkissingloopinteractions |