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Single bacterial resolvases first exploit, then constrain intrinsic dynamics of the Holliday junction to direct recombination

Homologous recombination forms and resolves an entangled DNA Holliday Junction (HJ) crucial for achieving genetic reshuffling and genome repair. To maintain genomic integrity, specialized resolvase enzymes cleave the entangled DNA into two discrete DNA molecules. However, it is unclear how two simil...

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Detalles Bibliográficos
Autores principales: Ray, Sujay, Pal, Nibedita, Walter, Nils G
Formato: Online Artículo Texto
Lenguaje:English
Publicado: Oxford University Press 2021
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC7969024/
https://www.ncbi.nlm.nih.gov/pubmed/33619520
http://dx.doi.org/10.1093/nar/gkab096
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author Ray, Sujay
Pal, Nibedita
Walter, Nils G
author_facet Ray, Sujay
Pal, Nibedita
Walter, Nils G
author_sort Ray, Sujay
collection PubMed
description Homologous recombination forms and resolves an entangled DNA Holliday Junction (HJ) crucial for achieving genetic reshuffling and genome repair. To maintain genomic integrity, specialized resolvase enzymes cleave the entangled DNA into two discrete DNA molecules. However, it is unclear how two similar stacking isomers are distinguished, and how a cognate sequence is found and recognized to achieve accurate recombination. We here use single-molecule fluorescence observation and cluster analysis to examine how prototypic bacterial resolvase RuvC singles out two of the four HJ strands and achieves sequence-specific cleavage. We find that RuvC first exploits, then constrains the dynamics of intrinsic HJ isomer exchange at a sampled branch position to direct cleavage toward the catalytically competent HJ conformation and sequence, thus controlling recombination output at minimal energetic cost. Our model of rapid DNA scanning followed by ‘snap-locking’ of a cognate sequence is strikingly consistent with the conformational proofreading of other DNA-modifying enzymes.
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spelling pubmed-79690242021-03-22 Single bacterial resolvases first exploit, then constrain intrinsic dynamics of the Holliday junction to direct recombination Ray, Sujay Pal, Nibedita Walter, Nils G Nucleic Acids Res Nucleic Acid Enzymes Homologous recombination forms and resolves an entangled DNA Holliday Junction (HJ) crucial for achieving genetic reshuffling and genome repair. To maintain genomic integrity, specialized resolvase enzymes cleave the entangled DNA into two discrete DNA molecules. However, it is unclear how two similar stacking isomers are distinguished, and how a cognate sequence is found and recognized to achieve accurate recombination. We here use single-molecule fluorescence observation and cluster analysis to examine how prototypic bacterial resolvase RuvC singles out two of the four HJ strands and achieves sequence-specific cleavage. We find that RuvC first exploits, then constrains the dynamics of intrinsic HJ isomer exchange at a sampled branch position to direct cleavage toward the catalytically competent HJ conformation and sequence, thus controlling recombination output at minimal energetic cost. Our model of rapid DNA scanning followed by ‘snap-locking’ of a cognate sequence is strikingly consistent with the conformational proofreading of other DNA-modifying enzymes. Oxford University Press 2021-02-22 /pmc/articles/PMC7969024/ /pubmed/33619520 http://dx.doi.org/10.1093/nar/gkab096 Text en © The Author(s) 2021. 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-NonCommercial 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 Nucleic Acid Enzymes
Ray, Sujay
Pal, Nibedita
Walter, Nils G
Single bacterial resolvases first exploit, then constrain intrinsic dynamics of the Holliday junction to direct recombination
title Single bacterial resolvases first exploit, then constrain intrinsic dynamics of the Holliday junction to direct recombination
title_full Single bacterial resolvases first exploit, then constrain intrinsic dynamics of the Holliday junction to direct recombination
title_fullStr Single bacterial resolvases first exploit, then constrain intrinsic dynamics of the Holliday junction to direct recombination
title_full_unstemmed Single bacterial resolvases first exploit, then constrain intrinsic dynamics of the Holliday junction to direct recombination
title_short Single bacterial resolvases first exploit, then constrain intrinsic dynamics of the Holliday junction to direct recombination
title_sort single bacterial resolvases first exploit, then constrain intrinsic dynamics of the holliday junction to direct recombination
topic Nucleic Acid Enzymes
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC7969024/
https://www.ncbi.nlm.nih.gov/pubmed/33619520
http://dx.doi.org/10.1093/nar/gkab096
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