Dna2 nuclease-helicase structure, mechanism and regulation by Rpa

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Cite this article as: eLife 2015;4:e09832 doi: 10.7554/eLife.09832

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Abstract

The Dna2 nuclease-helicase maintains genomic integrity by processing DNA double-strand breaks, Okazaki fragments and stalled replication forks. Dna2 requires ssDNA ends, and is dependent on the ssDNA-binding protein Rpa, which controls cleavage polarity. Here we present the 2.3 Å structure of intact mouse Dna2 bound to a 15-nucleotide ssDNA. The nuclease active site is embedded in a long, narrow tunnel through which the DNA has to thread. The helicase domain is required for DNA binding but not threading. We also present the structure of a flexibly-tethered Dna2-Rpa interaction that recruits Dna2 to Rpa-coated DNA. We establish that a second Dna2-Rpa interaction is mutually exclusive with Rpa-DNA interactions and mediates the displacement of Rpa from ssDNA. This interaction occurs at the nuclease tunnel entrance and the 5' end of the Rpa-DNA complex. Hence, it only displaces Rpa from the 5' but not 3' end, explaining how Rpa regulates cleavage polarity.

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Author details

Chun Zhou

Structural Biology Program, Memorial Sloan Kettering Cancer Center, New York, United States

Competing interests
The authors declare that no competing interests exist.
Sergei Pourmal

Structural Biology Program, Memorial Sloan Kettering Cancer Center, New York, United States

Competing interests
The authors declare that no competing interests exist.
Nikola P Pavletich

Structural Biology Program, Memorial Sloan Kettering Cancer Center, New York, United States

For correspondence
pavletin@mskcc.org

Competing interests
The authors declare that no competing interests exist.

Reviewing Editor

John Kuriyan, Howard Hughes Medical Institute, University of California, Berkeley, United States

Publication history

Received: July 2, 2015
Accepted: October 20, 2015
Accepted Manuscript published: October 22, 2015 (version 1)
Accepted Manuscript updated: November 2, 2015 (version 2)
Version of Record published: December 30, 2015 (version 3)

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This article is distributed under the terms of the Creative Commons Attribution License permitting unrestricted use and redistribution provided that the original author and source are credited.