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Structural transitions upon guide RNA binding and their importance in Cas12g-mediated RNA cleavage
Cas12g is an endonuclease belonging to the type V RNA-guided CRISPR–Cas family. It is known for its ability to cleave RNA substrates using a conserved endonuclease active site located in the RuvC domain. In this study, we determined the crystal structure of apo-Cas12g, the cryo-EM structure of the C...
Autores principales: | , , , , , , , , |
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Formato: | Online Artículo Texto |
Lenguaje: | English |
Publicado: |
Public Library of Science
2023
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Materias: | |
Acceso en línea: | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC10511118/ https://www.ncbi.nlm.nih.gov/pubmed/37729124 http://dx.doi.org/10.1371/journal.pgen.1010930 |
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author | Liu, Mengxi Li, Zekai Chen, Jing Lin, Jinying Lu, Qiuhua Ye, Yangmiao Zhang, Hongmin Zhang, Bo Ouyang, Songying |
author_facet | Liu, Mengxi Li, Zekai Chen, Jing Lin, Jinying Lu, Qiuhua Ye, Yangmiao Zhang, Hongmin Zhang, Bo Ouyang, Songying |
author_sort | Liu, Mengxi |
collection | PubMed |
description | Cas12g is an endonuclease belonging to the type V RNA-guided CRISPR–Cas family. It is known for its ability to cleave RNA substrates using a conserved endonuclease active site located in the RuvC domain. In this study, we determined the crystal structure of apo-Cas12g, the cryo-EM structure of the Cas12g-sgRNA binary complex and investigated conformational changes that occur during the transition from the apo state to the Cas12g-sgRNA binary complex. The conserved zinc finger motifs in Cas12g undergo an ordered-to-disordered transition from the apo to the sgRNA-bound state and their mutations negatively impact on target RNA cleavage. Moreover, we identified a lid motif in the RuvC domain that undergoes transformation from a helix to loop to regulate the access to the RuvC active site and subsequent cleavage of the RNA substrate. Overall, our study provides valuable insights into the mechanisms by which Cas12g recognizes sgRNA and the conformational changes it undergoes from sgRNA binding to the activation of the RNase active site, thereby laying a foundation for the potential repurposing of Cas12g as a tool for RNA-editing. |
format | Online Article Text |
id | pubmed-10511118 |
institution | National Center for Biotechnology Information |
language | English |
publishDate | 2023 |
publisher | Public Library of Science |
record_format | MEDLINE/PubMed |
spelling | pubmed-105111182023-09-21 Structural transitions upon guide RNA binding and their importance in Cas12g-mediated RNA cleavage Liu, Mengxi Li, Zekai Chen, Jing Lin, Jinying Lu, Qiuhua Ye, Yangmiao Zhang, Hongmin Zhang, Bo Ouyang, Songying PLoS Genet Research Article Cas12g is an endonuclease belonging to the type V RNA-guided CRISPR–Cas family. It is known for its ability to cleave RNA substrates using a conserved endonuclease active site located in the RuvC domain. In this study, we determined the crystal structure of apo-Cas12g, the cryo-EM structure of the Cas12g-sgRNA binary complex and investigated conformational changes that occur during the transition from the apo state to the Cas12g-sgRNA binary complex. The conserved zinc finger motifs in Cas12g undergo an ordered-to-disordered transition from the apo to the sgRNA-bound state and their mutations negatively impact on target RNA cleavage. Moreover, we identified a lid motif in the RuvC domain that undergoes transformation from a helix to loop to regulate the access to the RuvC active site and subsequent cleavage of the RNA substrate. Overall, our study provides valuable insights into the mechanisms by which Cas12g recognizes sgRNA and the conformational changes it undergoes from sgRNA binding to the activation of the RNase active site, thereby laying a foundation for the potential repurposing of Cas12g as a tool for RNA-editing. Public Library of Science 2023-09-20 /pmc/articles/PMC10511118/ /pubmed/37729124 http://dx.doi.org/10.1371/journal.pgen.1010930 Text en © 2023 Liu et al 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 use, distribution, and reproduction in any medium, provided the original author and source are credited. |
spellingShingle | Research Article Liu, Mengxi Li, Zekai Chen, Jing Lin, Jinying Lu, Qiuhua Ye, Yangmiao Zhang, Hongmin Zhang, Bo Ouyang, Songying Structural transitions upon guide RNA binding and their importance in Cas12g-mediated RNA cleavage |
title | Structural transitions upon guide RNA binding and their importance in Cas12g-mediated RNA cleavage |
title_full | Structural transitions upon guide RNA binding and their importance in Cas12g-mediated RNA cleavage |
title_fullStr | Structural transitions upon guide RNA binding and their importance in Cas12g-mediated RNA cleavage |
title_full_unstemmed | Structural transitions upon guide RNA binding and their importance in Cas12g-mediated RNA cleavage |
title_short | Structural transitions upon guide RNA binding and their importance in Cas12g-mediated RNA cleavage |
title_sort | structural transitions upon guide rna binding and their importance in cas12g-mediated rna cleavage |
topic | Research Article |
url | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC10511118/ https://www.ncbi.nlm.nih.gov/pubmed/37729124 http://dx.doi.org/10.1371/journal.pgen.1010930 |
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