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PMID:21293191

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Citation

Roy, MA, Siddiqui, N and D'Amours, D (2011) Dynamic and selective DNA-binding activity of Smc5, a core component of the Smc5-Smc6 complex. Cell Cycle 10:690-700

Abstract

Members of the structural maintenance of chromosome (SMC) family of proteins are essential regulators of genomic stability. In particular, the conserved Smc5-6 complex is required for efficient DNA repair, checkpoint signaling, and DNA replication in all eukaryotes. Despite these important functions, the actual nature of the DNA substrates recognized by the Smc5-6 complex in chromosomes is currently unknown. Furthermore, how the core SMC components of the Smc5-6 complex use their ATPase-driven mechanochemical activities to act on chromosomes is not understood. Here, we address these issues by purifying and defining the DNA-binding activity of Smc5. We show that Smc5 binds strongly and specifically to single-stranded DNA (ssDNA). Remarkably, this DNA-binding activity is independent of Smc6 and is observed with the monomeric form of Smc5. We further show that Smc5 ATPase activity is essential for its functions in vivo and that ATP regulates the association of Smc5 with its substrates in vitro. Finally, we demonstrate that Smc5 is able to bind efficiently to oligonucleotides consistent in size with ssDNA intermediates produced during DNA replication and repair. Collectively, our data on the DNA-binding activities of Smc5 provide a compelling molecular basis for the role of the Smc5-6 complex in the DNA damage response.

Links

PubMed

Keywords

Adenosine Triphosphatases/metabolism; Adenosine Triphosphate/metabolism; Cell Cycle; Cell Cycle Proteins/isolation & purification; Cell Cycle Proteins/metabolism; Chromosomes/chemistry; DNA Repair; DNA, Fungal/genetics; DNA, Fungal/metabolism; DNA, Single-Stranded/metabolism; DNA-Binding Proteins/metabolism; Protein Binding; Saccharomyces cerevisiae/enzymology; Saccharomyces cerevisiae/genetics; Saccharomyces cerevisiae/metabolism; Saccharomyces cerevisiae Proteins/isolation & purification; Saccharomyces cerevisiae Proteins/metabolism

Significance

Annotations

Gene product Qualifier GO Term Evidence Code with/from Aspect Extension Notes Status

YEAST:SMC5

enables

GO:0016887: ATP hydrolysis activity

ECO:0000314: direct assay evidence used in manual assertion

F

Seeded From UniProt

complete

YEAST:SMC5

enables

GO:0003697: single-stranded DNA binding

ECO:0000314: direct assay evidence used in manual assertion

F

Seeded From UniProt

complete


See also

References

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