Drf1-dependent kinase interacts with claspin through a conserved protein motif

Daniel Gold, William G. Dunphy

Research output: Contribution to journalArticle

2 Citations (Scopus)

Abstract

The Dbf4/Drf1-dependent kinase (DDK) is required for the initiation of DNA replication in eukaryotes. Another protein, Claspin, mediates the activation of a cellular checkpoint response to stalled replication forks and is also a regulator of replication. In this study, we found that DDK phosphorylates Claspin in vitro and forms a nuclear complex containing Cdc7, Drf1, and Claspin in Xenopus egg extracts. In addition, purified Claspin and DDK are capable of a direct in vitro interaction.We identified a conserved binding site on Claspin required for its interaction with DDK. This site corresponds to the first of two sequence repeats in the Chk1-binding domain of Claspin. Furthermore, we have established that two amino acids in this motif, Asp861 and Gln 866, are essential for the interaction between Claspin and DDK. We found that mutant forms of Claspin incapable of interacting with DDK are still able to associate with and activate Chk1 in response to DNA replication blockages. However, Claspin-depleted egg extracts that have been reconstituted with these mutants of Claspin undergo DNA replication more slowly. These findings suggest that the interaction of DDK with Claspin mediates a checkpoint-independent function of Claspin related to DNA replication.

Original languageEnglish (US)
Pages (from-to)12638-12646
Number of pages9
JournalJournal of Biological Chemistry
Volume285
Issue number17
DOIs
StatePublished - Apr 23 2010
Externally publishedYes

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Amino Acid Motifs
Phosphotransferases
DNA Replication
Proteins
DNA
Ovum
Eukaryota
Chemical activation
Binding Sites
Amino Acids

ASJC Scopus subject areas

  • Biochemistry
  • Cell Biology
  • Molecular Biology

Cite this

Drf1-dependent kinase interacts with claspin through a conserved protein motif. / Gold, Daniel; Dunphy, William G.

In: Journal of Biological Chemistry, Vol. 285, No. 17, 23.04.2010, p. 12638-12646.

Research output: Contribution to journalArticle

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