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Crystal structure of a bacteriophage T7 DNA replication complex at 2.2 Å resolution

Nature volume 391, pages 251–258 (1998)Cite this article

Abstract

DNA polymerases change their specificity for nucleotide substrates with each catalytic cycle, while achieving error frequencies in the range of 10−5to 10−6. Here we present a 2.2 Å crystal structure of the replicative DNA polymerase from bacteriophage T7 complexed with a primer–template and a nucleoside triphosphate in the polymerase active site. The structure illustrates how nucleotides are selected in a template-directed manner, and provides a structural basis for a metal-assisted mechanism of phosphoryl transfer by a large group of related polymerases.

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Figure 1: Topology and overall fold of T7 DNA polymerase.
Figure 2: Polymerase contacts to the primer–template.
Figure 3: Two-metal ligation of the bound nucleotide.
Figure 4: Selectivity for 2′-deoxyribonucleotides.
Figure 5: Open and closed conformations of polymerase active sites.
Figure 6: Misincorporation and proofreading.

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Acknowledgements

We thank R. M. Sweet and F. W. Studier for access and assistance with the use of beamline X12C (NSLS, Brookhaven National Laboratory). We thank H. J. Kwon, A. Lau and M. Rould for experimental help at the synchrotron, members of the Ellenberger, Richardson and Hogle groups for many discussions and M. Rould for programs and discussions. This work was supported by the National Institutes of Health, the Lucille Markey Charitable Trust, the Department of Energy, the Harvard Center for Structural Biology and the Armenise-Harvard Foundation for Advanced Scientific Research. C.C.R. and S.T. are consultants to Amersham Life Science Inc., which has licenses from Harvard University to commercialize DNA polymerases for use in DNA sequencing.

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  1. Department of Biological Chemistry and Molecular Pharmacology, Harvard Medical School, 240 Longwood Avenue, Boston, 02115, Massachusetts, USA

    Sylvie Doublié, Stanley Tabor, Alexander M. Long, Charles C. Richardson & Tom Ellenberger

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Correspondence to Tom Ellenberger.

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Doublié, S., Tabor, S., Long, A. et al. Crystal structure of a bacteriophage T7 DNA replication complex at 2.2 Å resolution. Nature 391, 251–258 (1998). https://doi.org/10.1038/34593

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