Structural mechanism for the selective phosphorylation of DNA-loaded MCM double hexamers by the Dbf4-dependent kinase.

Autor: Greiwe JF; Macromolecular Machines Laboratory, The Francis Crick Institute, London, UK., Miller TCR; Macromolecular Machines Laboratory, The Francis Crick Institute, London, UK.; Center for Chromosome Stability, University of Copenhagen, Copenhagen, Denmark., Locke J; Macromolecular Machines Laboratory, The Francis Crick Institute, London, UK., Martino F; Macromolecular Machines Laboratory, The Francis Crick Institute, London, UK.; Human Technopole, Milan, Italy., Howell S; Proteomics Science Technology Platform, The Francis Crick Institute, London, UK., Schreiber A; Cellular Degradation Systems Laboratory, The Francis Crick Institute, London, UK., Nans A; Structural Biology Science Technology Platform, The Francis Crick Institute, London, UK., Diffley JFX; Chromosome Replication Laboratory, The Francis Crick Institute, London, UK., Costa A; Macromolecular Machines Laboratory, The Francis Crick Institute, London, UK. alessandro.costa@crick.ac.uk.
Jazyk: angličtina
Zdroj: Nature structural & molecular biology [Nat Struct Mol Biol] 2022 Jan; Vol. 29 (1), pp. 10-20. Date of Electronic Publication: 2021 Dec 28.
DOI: 10.1038/s41594-021-00698-z
Abstrakt: Loading of the eukaryotic replicative helicase onto replication origins involves two MCM hexamers forming a double hexamer (DH) around duplex DNA. During S phase, helicase activation requires MCM phosphorylation by Dbf4-dependent kinase (DDK), comprising Cdc7 and Dbf4. DDK selectively phosphorylates loaded DHs, but how such fidelity is achieved is unknown. Here, we determine the cryogenic electron microscopy structure of Saccharomyces cerevisiae DDK in the act of phosphorylating a DH. DDK docks onto one MCM ring and phosphorylates the opposed ring. Truncation of the Dbf4 docking domain abrogates DH phosphorylation, yet Cdc7 kinase activity is unaffected. Late origin firing is blocked in response to DNA damage via Dbf4 phosphorylation by the Rad53 checkpoint kinase. DDK phosphorylation by Rad53 impairs DH phosphorylation by blockage of DDK binding to DHs, and also interferes with the Cdc7 active site. Our results explain the structural basis and regulation of the selective phosphorylation of DNA-loaded MCM DHs, which supports bidirectional replication.
(© 2021. The Author(s).)
Databáze: MEDLINE
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