Structure of human Cdc45 and implications for CMG helicase function.

Aline C Simon,Vincenzo Sannino,Vincenzo Costanzo,Luca Pellegrini

doi:10.1038/ncomms11638

Aline C Simon, Vincenzo Sannino + Show 2 more

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https://doi.org/10.1038/ncomms11638

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Abstract

Cell division cycle protein 45 (Cdc45) is required for DNA synthesis during genome duplication, as a component of the Cdc45-MCM-GINS (CMG) helicase. Despite its essential biological function, its biochemical role in DNA replication has remained elusive. Here we report the 2.1-Å crystal structure of human Cdc45, which confirms its evolutionary link with the bacterial RecJ nuclease and reveals several unexpected features that underpin its function in eukaryotic DNA replication. These include a long-range interaction between N- and C-terminal DHH domains, blocking access to the DNA-binding groove of its RecJ-like fold, and a helical insertion in its N-terminal DHH domain, which appears poised for replisome interactions. In combination with available electron microscopy data, we validate by mutational analysis the mechanism of Cdc45 association with the MCM ring and GINS co-activator, critical for CMG assembly. These findings provide an indispensable molecular basis to rationalize the essential role of Cdc45 in genomic duplication.

Highlights

Cell division cycle protein 45 (Cdc45) is required for DNA synthesis during genome duplication, as a component of the Cdc45-MCM-GINS (CMG) helicase
Experimental work spanning several decades in model systems such as yeast has led to the identification of a complete list of protein factors necessary for DNA synthesis in vitro
Extensive experimental evidence has since demonstrated that cell division cycle protein 45 (Cdc45) is required throughout the process of DNA replication: it is essential for establishment of an initiation complex at DNA origins[6], and for chromosome unwinding and DNA synthesis at replication forks[7,8]

Summary

Introduction

Cell division cycle protein 45 (Cdc45) is required for DNA synthesis during genome duplication, as a component of the Cdc45-MCM-GINS (CMG) helicase. In combination with available electron microscopy data, we validate by mutational analysis the mechanism of Cdc[45] association with the MCM ring and GINS co-activator, critical for CMG assembly These findings provide an indispensable molecular basis to rationalize the essential role of Cdc[45] in genomic duplication. Extensive experimental evidence has since demonstrated that cell division cycle protein 45 (Cdc45) is required throughout the process of DNA replication: it is essential for establishment of an initiation complex at DNA origins[6], and for chromosome unwinding and DNA synthesis at replication forks[7,8]. Association of replication factors MCM10 and Ctf[4] with the CMG leads to recruitment of RPA and DNA polymerase a/primase, with consequent initiation of DNA synthesis[21,22]

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