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Biochemical analysis of human Dna2

Masuda-Sasa, Taro and Imamura, Osamu and Campbell, Judith L. (2006) Biochemical analysis of human Dna2. Nucleic Acids Research, 34 (6). pp. 1865-1875. ISSN 0305-1048. doi:10.1093/nar/gkl070.

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Yeast Dna2 helicase/nuclease is essential for DNA replication and assists FEN1 nuclease in processing a subset of Okazaki fragments that have long single-stranded 5' flaps. It is also involved in the maintenance of telomeres. DNA2 is a gene conserved in eukaryotes, and a putative human ortholog of yeast DNA2 (ScDNA2) has been identified. Little is known about the role of human DNA2 (hDNA2), although complementation experiments have shown that it can function in yeast to replace ScDNA2. We have now characterized the biochemical properties of hDna2. Recombinant hDna2 has single-stranded DNA-dependent ATPase and DNA helicase activity. It also has 5'–3' nuclease activity with preference for single-stranded 5' flaps adjacent to a duplex DNA region. The nuclease activity is stimulated by RPA and suppressed by steric hindrance at the 5' end. Moreover, hDna2 shows strong 3'–5' nuclease activity. This activity cleaves single-stranded DNA in a fork structure and, like the 5'–3' activity, is suppressed by steric hindrance at the 3'-end, suggesting that the 3'–5' nuclease requires a 3' single-stranded end for activation. These biochemical specificities are very similar to those of the ScDna2 protein, but suggest that the 3'–5' nuclease activity may be more important than previously thought.

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Additional Information:© The Author 2006. Published by Oxford University Press. Received February 2, 2006. Revised March 2, 2006. Accepted March 2, 2006. We thank Dr Peter Snow of the Caltech Protein Expression Center for expression and purification of hDNA2. We thank Dr Carl Parker and Dr Marc Wold for providing necessary reagents. Funding for this research and funding to pay the Open Access publication charges for this article were provided by USPHS GM25508, and grants from the Margaret E. Early Trust and the Research Management Group. Conflict of interest statement. None declared. The online version of this article has been published under an open access model. Users are entitled to use, reproduce, disseminate, or display the open access version of this article for non-commercial purposes provided that: the original authorship is properly and fully attributed; the Journal and Oxford University Press are attributed as the original place of publication with the correct citation details given; if an article is subsequently reproduced or disseminated not in its entirety but only in part or as a derivative work this must be clearly indicated.
Issue or Number:6
Record Number:CaltechAUTHORS:MASnar06
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Usage Policy:No commercial reproduction, distribution, display or performance rights in this work are provided.
ID Code:2529
Deposited By: Archive Administrator
Deposited On:07 Apr 2006
Last Modified:08 Nov 2021 19:48

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