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Cell Rep. 2014 Mar 27;6(6):1129-1138. doi: 10.1016/j.celrep.2014.02.025. Epub 2014 Mar 13.

Helicase and polymerase move together close to the fork junction and copy DNA in one-nucleotide steps.

Author information

1
Department of Biochemistry and Molecular Biology, Rutgers, The State University of New Jersey, Robert Wood Johnson Medical School, Piscataway, NJ 08854, USA.
2
Department of Biochemistry and Molecular Biology, Rutgers, The State University of New Jersey, Robert Wood Johnson Medical School, Piscataway, NJ 08854, USA. Electronic address: patelss@rwjms.rutgers.edu.

Abstract

By simultaneously measuring DNA synthesis and dNTP hydrolysis, we show that T7 DNA polymerase and T7 gp4 helicase move in sync during leading-strand synthesis, taking one-nucleotide steps and hydrolyzing one dNTP per base-pair unwound/copied. The cooperative catalysis enables the helicase and polymerase to move at a uniformly fast rate without guanine:cytosine (GC) dependency or idling with futile NTP hydrolysis. We show that the helicase and polymerase are located close to the replication fork junction. This architecture enables the polymerase to use its strand-displacement synthesis to increase the unwinding rate, whereas the helicase aids this process by translocating along single-stranded DNA and trapping the unwound bases. Thus, in contrast to the helicase-only unwinding model, our results suggest a model in which the helicase and polymerase are moving in one-nucleotide steps, DNA synthesis drives fork unwinding, and a role of the helicase is to trap the unwound bases and prevent DNA reannealing.

PMID:
24630996
PMCID:
PMC4010093
DOI:
10.1016/j.celrep.2014.02.025
[Indexed for MEDLINE]
Free PMC Article

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