cryo-EM structures of the E. coli replicative DNA polymerase reveal dynamic interactions with clamp, exonuclease and τ

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Abstract

The replicative DNA polymerase PolIIIα from E. coli is a uniquely fast and processive enzyme. For its activity it relies on the DNA sliding clamp β, the proofreading exonuclease ε and the C-terminal domain of the clamp loader subunit τ. Due to the dynamic nature of the four-protein complex it has long been refractory to structural characterization. Here we present the 8 Å resolution cryo-electron microscopy structures of DNA-bound and DNA-free states of the PolIII-clamp-exonuclease-τc complex. The structures show how the polymerase is tethered to the DNA through multiple contacts with the clamp and exonuclease. A novel contact between the polymerase and clamp is made in the DNA bound state, facilitated by a large movement of the polymerase tail domain and τc. These structures provide crucial insights into the organization of the catalytic core of the replisome and form an important step towards determining the structure of the complete holoenzyme.

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Rafael Fernández-Leiro

Structural Studies, MRC laboratory of Molecular Biology, Cambridge, United Kingdom

Competing interests
No competing interests declared.
Julian Conrad

Structural Studies, MRC laboratory of Molecular Biology, Cambridge, United Kingdom

Competing interests
No competing interests declared.
Sjors HW Scheres

Structural Studies, Medical Research Council Laboratory of Molecular Biology, Cambridge, United Kingdom

Competing interests
Sjors HW Scheres, Reviewing editor, eLife.
Meindert Hugo Lamers

Structural Studies, Medical Research Council Laboratory of Molecular Biology, Cambridge, United Kingdom

For correspondence
mlamers@mrc-lmb.cam.ac.uk

Competing interests
No competing interests declared.

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