Dataset Information

Archaeal orthologs of Cdc45 and GINS form a stable complex that stimulates the helicase activity of MCM.

ABSTRACT: The regulated recruitment of Cdc45 and GINS is key to activating the eukaryotic MCM(2-7) replicative helicase. We demonstrate that the homohexameric archaeal MCM helicase associates with orthologs of GINS and Cdc45 in vivo and in vitro. Association of these factors with MCM robustly stimulates the MCM helicase activity. In contrast to the situation in eukaryotes, archaeal Cdc45 and GINS form an extremely stable complex before binding MCM. Further, the archaeal GINS•Cdc45 complex contains two copies of Cdc45. Our analyses give insight into the function and evolution of the conserved core of the archaeal/eukaryotic replisome.

SUBMITTER: Xu Y

PROVIDER: S-EPMC5127375 | biostudies-literature | 2016 Nov

REPOSITORIES: biostudies-literature

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Archaeal orthologs of Cdc45 and GINS form a stable complex that stimulates the helicase activity of MCM.

Xu Yuli Y Gristwood Tamzin T Hodgson Ben B Trinidad Jonathan C JC Albers Sonja-Verena SV Bell Stephen D SD

Proceedings of the National Academy of Sciences of the United States of America 20161107 47

The regulated recruitment of Cdc45 and GINS is key to activating the eukaryotic MCM(2-7) replicative helicase. We demonstrate that the homohexameric archaeal MCM helicase associates with orthologs of GINS and Cdc45 in vivo and in vitro. Association of these factors with MCM robustly stimulates the MCM helicase activity. In contrast to the situation in eukaryotes, archaeal Cdc45 and GINS form an extremely stable complex before binding MCM. Further, the archaeal GINS•Cdc45 complex contains two cop ...[more]

PMID: 27821767

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Project description:UnlabelledGINS is a key component of the eukaryotic Cdc45-minichromosome maintenance (MCM)-GINS (CMG) complex, which unwinds duplex DNA at the moving replication fork. Archaeal GINS complexes have been shown to stimulate the helicase activity of their cognate MCM mainly by elevating its ATPase activity. Here, we report that GINS from the thermoacidophilic crenarchaeon Sulfolobus solfataricus (SsoGINS) is capable of DNA binding and binds preferentially to single-stranded DNA (ssDNA) over double-stranded DNA (dsDNA). Notably, SsoGINS binds more strongly to dsDNA with a 5' ssDNA tail than to dsDNA with a 3' tail and more strongly to an ssDNA fragment blocked at the 3' end than to one at the 5' end with a biotin-streptavidin (SA) complex, suggesting the ability of the protein complex to slide in a 5'-to-3' direction along ssDNA. DNA-bound SsoGINS enhances DNA binding by SsoMCM. Furthermore, SsoGINS increases the helicase activity of SsoMCM. However, the ATPase activity of SsoMCM is not affected by SsoGINS. Our results suggest that SsoGINS facilitates processive DNA unwinding by SsoMCM by enhancing the binding of the helicase to DNA. We propose that SsoGINS stabilizes the interaction of SsoMCM with the replication fork and moves along with the helicase as the fork progresses.ImportanceGINS is a key component of the eukaryotic Cdc45-MCM-GINS complex, a molecular motor that drives the unwinding of DNA in front of the replication fork. Archaea also encode GINS, which interacts with MCM, the helicase. But how archaeal GINS serves its role remains to be understood. In this study, we show that GINS from the hyperthermophilic archaeon Sulfolobus solfataricus is able to bind to DNA and slide along ssDNA in a 5'-to-3' direction. Furthermore, Sulfolobus GINS enhances DNA binding by MCM, which slides along ssDNA in a 3'-to-5' direction. Taken together, these results suggest that Sulfolobus GINS may stabilize the interaction of MCM with the moving replication fork, facilitating processive DNA unwinding.

Dataset Information

Archaeal orthologs of Cdc45 and GINS form a stable complex that stimulates the helicase activity of MCM.

Publications

Archaeal orthologs of Cdc45 and GINS form a stable complex that stimulates the helicase activity of MCM.

Similar Datasets

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