Mechanism of opening a sliding Clamp

Lauren G. Douma, Kevin K. Yu, Jennifer K. England, Marcia Levitus, Linda B. Bloom

Research output: Contribution to journalArticle

4 Scopus citations

Abstract

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-openlock' 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.

Original languageEnglish (US)
Pages (from-to)10178-10189
Number of pages12
JournalNucleic acids research
Volume45
Issue number17
DOIs
StatePublished - Sep 1 2017

ASJC Scopus subject areas

  • Genetics

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    Douma, L. G., Yu, K. K., England, J. K., Levitus, M., & Bloom, L. B. (2017). Mechanism of opening a sliding Clamp. Nucleic acids research, 45(17), 10178-10189. https://doi.org/10.1093/nar/gkx665