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Dna2 Exhibits a Unique Strand End-dependent Helicase Function

Balakrishnan, Lata and Polaczek, Piotr and Pokharel, Subhash and Campbell, Judith L. and Bambara, Robert A. (2010) Dna2 Exhibits a Unique Strand End-dependent Helicase Function. Journal of Biological Chemistry, 285 (50). pp. 38861-38868. ISSN 0021-9258. PMCID PMC2998112.

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Dna2 endonuclease/helicase participates in eukaryotic DNA transactions including cleavage of long flaps generated during Okazaki fragment processing. Its unusual substrate interaction consists of recognition and binding of the flap base, then threading over the 5′-end of the flap, and cleaving periodically to produce a terminal product ~5 nt in length. Blocking the 5′-end prevents cleavage. The Dna2 ATP-driven 5′ to 3′ DNA helicase function promotes motion of Dna2 on the flap, presumably aiding its nuclease function. Here we demonstrate using two different nuclease-dead Dna2 mutants that on substrates simulating Okazaki fragments, Dna2 must thread onto an unblocked 5′ flap to display helicase activity. This requirement is maintained on substrates with single-stranded regions thousands of nucleotides in length. To our knowledge this is the first description of a eukaryotic helicase that cannot load onto its tracking strand internally but instead must enter from the end. Biologically, the loading requirement likely helps the helicase to coordinate with the Dna2 nuclease function to prevent creation of undesirably long flaps during DNA transactions.

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Additional Information:© 2010 The American Society for Biochemistry and Molecular Biology, Inc. Received July 19, 2010. Revision received September 12, 2010. This work was supported, in whole or in part, by National Institutes of Health Grants GM024441 (to R. A. B.) and GM087666 (to J. L. C.). We thank members of the Bambara and Campbell laboratories for helpful suggestions and discussions. Special thanks to Dr. Steven Matson for critical reading of our manuscript. We would also like to thank Dr. Robert Brosh for critical discussions about the helicase activity.
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Subject Keywords:DNA Enzymes, DNA Helicase, DNA Repair, DNA Replication, Enzyme Mechanisms, Dna2, Okazaki Fragment Maturation
Issue or Number:50
PubMed Central ID:PMC2998112
Record Number:CaltechAUTHORS:20110103-095820565
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Official Citation:Balakrishnan, L., P. Polaczek, et al. (2010). "Dna2 Exhibits a Unique Strand End-dependent Helicase Function." Journal of Biological Chemistry 285(50): 38861-38868.
Usage Policy:No commercial reproduction, distribution, display or performance rights in this work are provided.
ID Code:21530
Deposited By: Tony Diaz
Deposited On:25 Jan 2011 22:06
Last Modified:03 Oct 2019 02:25

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