Journal
PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA
Volume 114, Issue 16, Pages 4141-4146Publisher
NATL ACAD SCIENCES
DOI: 10.1073/pnas.1700530114
Keywords
DNA replication; CMG helicase; DNA polymerase; single-particle electron microscopy
Categories
Funding
- Francis Crick Institute from Cancer Research UK
- UK Medical Research Council
- Wellcome Trust (WT)
- Boehringer Ingelheim Fonds
- European Research Council [669424-CHROMOREP]
- Wellcome Senior Investigator Award [106252/Z/14/Z]
- WT
- WT JIF Award [060208/Z/00/Z]
- WT [093305/Z/10/Z]
- Wellcome Trust [093305/Z/10/Z] Funding Source: Wellcome Trust
- Cancer Research UK [15669, 15852] Funding Source: researchfish
- Cancer Research UK
- The Francis Crick Institute [10065, 10066] Funding Source: researchfish
- The Francis Crick Institute [10349, 10397] Funding Source: researchfish
- Wellcome Trust [106252/Z/14/Z] Funding Source: researchfish
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The replisome unwinds and synthesizes DNA for genome duplication. In eukaryotes, the Cdc45-MCM-GINS (CMG) helicase and the leading-strand polymerase, Pol epsilon, form a stable assembly. The mechanism for coupling DNA unwinding with synthesis is starting to be elucidated, however the architecture and dynamics of the replication fork remain only partially understood, preventing a molecular understanding of chromosome replication. To address this issue, we conducted a systematic single-particle EM study on multiple permutations of the reconstituted CMG-Pol epsilon assembly. Pol epsilon contains two flexibly tethered lobes. The noncatalytic lobe is anchored to the motor of the helicase, whereas the polymerization domain extends toward the side of the helicase. We observe two alternate configurations of the DNA synthesis domain in the CMG-bound Pol epsilon. We propose that this conformational switch might control DNA template engagement and release, modulating replisome progression.
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