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Mechanism of opening a sliding clamp

Clamp loaders load ring-shaped sliding clamps onto DNA where the clamps serve as processivity factors for DNA polymerases. In the first stage of clamp loading, clamp loaders bind and stabilize clamps in an open conformation, and in the second stage, clamp loaders place the open clamps around DNA so...

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Autores principales: Douma, Lauren G., Yu, Kevin K., England, Jennifer K., Levitus, Marcia, Bloom, Linda B.
Formato: Online Artículo Texto
Lenguaje:English
Publicado: Oxford University Press 2017
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5737080/
https://www.ncbi.nlm.nih.gov/pubmed/28973453
http://dx.doi.org/10.1093/nar/gkx665
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author Douma, Lauren G.
Yu, Kevin K.
England, Jennifer K.
Levitus, Marcia
Bloom, Linda B.
author_facet Douma, Lauren G.
Yu, Kevin K.
England, Jennifer K.
Levitus, Marcia
Bloom, Linda B.
author_sort Douma, Lauren G.
collection PubMed
description Clamp loaders load ring-shaped sliding clamps onto DNA where the clamps serve as processivity factors for DNA polymerases. In the first stage of clamp loading, clamp loaders bind and stabilize clamps in an open conformation, and in the second stage, clamp loaders place the open clamps around DNA so that the clamps encircle DNA. Here, the mechanism of the initial clamp opening stage is investigated. Mutations were introduced into the Escherichia coli β-sliding clamp that destabilize the dimer interface to determine whether the formation of an open clamp loader–clamp complex is dependent on spontaneous clamp opening events. In other work, we showed that mutation of a positively charged Arg residue at the β-dimer interface and high NaCl concentrations destabilize the clamp, but neither facilitates the formation of an open clamp loader–clamp complex in experiments presented here. Clamp opening reactions could be fit to a minimal three-step ‘bind-open-lock’ model in which the clamp loader binds a closed clamp, the clamp opens, and subsequent conformational rearrangements ‘lock’ the clamp loader–clamp complex in a stable open conformation. Our results support a model in which the E. coli clamp loader actively opens the β-sliding clamp.
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spelling pubmed-57370802018-01-08 Mechanism of opening a sliding clamp Douma, Lauren G. Yu, Kevin K. England, Jennifer K. Levitus, Marcia Bloom, Linda B. Nucleic Acids Res Nucleic Acid Enzymes Clamp loaders load ring-shaped sliding clamps onto DNA where the clamps serve as processivity factors for DNA polymerases. In the first stage of clamp loading, clamp loaders bind and stabilize clamps in an open conformation, and in the second stage, clamp loaders place the open clamps around DNA so that the clamps encircle DNA. Here, the mechanism of the initial clamp opening stage is investigated. Mutations were introduced into the Escherichia coli β-sliding clamp that destabilize the dimer interface to determine whether the formation of an open clamp loader–clamp complex is dependent on spontaneous clamp opening events. In other work, we showed that mutation of a positively charged Arg residue at the β-dimer interface and high NaCl concentrations destabilize the clamp, but neither facilitates the formation of an open clamp loader–clamp complex in experiments presented here. Clamp opening reactions could be fit to a minimal three-step ‘bind-open-lock’ model in which the clamp loader binds a closed clamp, the clamp opens, and subsequent conformational rearrangements ‘lock’ the clamp loader–clamp complex in a stable open conformation. Our results support a model in which the E. coli clamp loader actively opens the β-sliding clamp. Oxford University Press 2017-09-29 2017-07-26 /pmc/articles/PMC5737080/ /pubmed/28973453 http://dx.doi.org/10.1093/nar/gkx665 Text en © The Author(s) 2017. 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 Nucleic Acid Enzymes
Douma, Lauren G.
Yu, Kevin K.
England, Jennifer K.
Levitus, Marcia
Bloom, Linda B.
Mechanism of opening a sliding clamp
title Mechanism of opening a sliding clamp
title_full Mechanism of opening a sliding clamp
title_fullStr Mechanism of opening a sliding clamp
title_full_unstemmed Mechanism of opening a sliding clamp
title_short Mechanism of opening a sliding clamp
title_sort mechanism of opening a sliding clamp
topic Nucleic Acid Enzymes
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5737080/
https://www.ncbi.nlm.nih.gov/pubmed/28973453
http://dx.doi.org/10.1093/nar/gkx665
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