cryo-EM structures of the <i>E. coli</i> replicative DNA polymerase reveal its dynamic interactions with the DNA sliding clamp, exonuclease and <b>τ</b>.

Fernandez-Leiro, Rafael; Conrad, Julian; Scheres, Sjors Hw; Lamers, Meindert H · Elife · 2015

basic_science · Level V

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Abstract

The replicative DNA polymerase PolIIIα from <i>Escherichia coli</i> 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-τ<sub>c</sub> 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 τ<sub>c</sub>. 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.