Structural Basis for DNA Binding by Replication Initiator Mcm10

Eric M. Warren, Sivaraja Vaithiyalingam, Justin Haworth, Briana Greer, Anja Katrin Bielinsky, Walter J. Chazin, Brandt F. Eichman

Research output: Contribution to journalArticlepeer-review

43 Scopus citations

Abstract

Mcm10 is an essential eukaryotic DNA replication protein required for assembly and progression of the replication fork. The highly conserved internal domain (Mcm10-ID) has been shown to physically interact with single-stranded (ss) DNA, DNA polymerase α, and proliferating cell nuclear antigen (PCNA). The crystal structure of Xenopus laevis Mcm10-ID presented here reveals a DNA binding architecture composed of an oligonucleotide/oligosaccharide-fold followed in tandem by a variant and highly basic zinc finger. NMR chemical shift perturbation and mutational studies of DNA binding activity in vitro reveal how Mcm10 uses this unique surface to engage ssDNA. Corresponding mutations in Saccharomyces cerevisiae result in increased sensitivity to replication stress, demonstrating the functional importance of DNA binding by this region of Mcm10 to replication. In addition, mapping Mcm10 mutations known to disrupt PCNA, polymerase α, and DNA interactions onto the crystal structure provides insight into how Mcm10 might coordinate protein and DNA binding within the replisome.

Original languageEnglish (US)
Pages (from-to)1892-1901
Number of pages10
JournalStructure
Volume16
Issue number12
DOIs
StatePublished - Dec 12 2008

Bibliographical note

Funding Information:
The authors are indebted to Audrey Metz for her assistance in crystal preparation, and to Johannes Walter for stimulating discussions. We thank the SER-CAT staff at the Advanced Photon Source (Argonne, IL), and Hassane Mchaourab and Albert Beth for access to fluorimeters. This work was funded by the National Institutes of Health (R01 GM080570 to B.F.E.; R01 GM065484 to W.J.C.; R01 GM074917 to A.-K.B.). Additional support for facilities came from the Vanderbilt Center in Molecular Toxicology (P30 ES000267) and the Vanderbilt Discovery Grant Program. E.M.W. was supported by the Molecular Biophysics Training Grant (T32 GM08320).

Keywords

  • DNA

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