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How a homolog of high-fidelity replicases conducts mutagenic DNA synthesis

Nature Structural & Molecular Biology volume 22, pages 298303 (2015) | Download Citation

Abstract

All DNA replicases achieve high fidelity by a conserved mechanism, but each translesion polymerase carries out mutagenic DNA synthesis in its own way. Here we report crystal structures of human DNA polymerase ν (Pol ν), which is homologous to high-fidelity replicases yet is error prone. Instead of a simple open-to-closed movement of the O helix upon binding of a correct incoming nucleotide, Pol ν has a different open state and requires the finger domain to swing sideways and undergo both opening and closing motions to accommodate the nascent base pair. A single–amino acid substitution in the O helix of the finger domain improves the fidelity of Pol ν nearly ten-fold. A unique cavity and the flexibility of the thumb domain allow Pol ν to generate and accommodate a looped-out primer strand. Primer loop-out may be a mechanism for DNA trinucloetide-repeat expansion.

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Acknowledgements

We thank D. Leahy and R. Craigie for editing the manuscript. The research was supported by the intramural research program of the US National Institutes of Health (DK036146-08, W.Y.).

Author information

Author notes

    • Young-Sam Lee

    Present address: Well-Aging Research Center, Samsung Advanced Institute of Technology, Suwon, Korea.

Affiliations

  1. Laboratory of Molecular Biology, National Institute of Diabetes and Digestive and Kidney Diseases, National Institutes of Health, Bethesda, Maryland, USA.

    • Young-Sam Lee
    • , Yang Gao
    •  & Wei Yang

Authors

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Contributions

Y.-S.L. carried out most of the experiments. Y.G. helped with kinetic measurement of K678A-mutant Pol ν and with data deposition. Y.-S.L. and W.Y. designed the project and prepared the manuscript.

Competing interests

The authors declare no competing financial interests.

Corresponding author

Correspondence to Wei Yang.

Integrated supplementary information

Supplementary information

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  1. 1.

    Supplementary Text and Figures

    Supplementary Figures 1–5 and Supplementary Tables 1 and 2

Videos

  1. 1.

    Finger movement between the conventional open and closed state exemplified by bacillus Pol I

    The binary complex (4BDP) and ternary complex (3THV) were compared. The three domains of the catalytic core are color-coded and the Exo domain is shown in silver. The M, N (lilac) and O (purple) helices undergo the large closing movement (40° rotation). The Oa and Ob helices move slightly towards the N and O helices.

  2. 2.

    Conformational changes of Pol ν from the unusual open (Ndna3) to the closed state (Ndna2).

    The Oa and Ob helices are colored in light blue. The remaining regions are colored as in movie 1. The M helix is disordered in Pol ν. The N (lilac) and O (purple) helices undergo a closing movement towards the DNA (25° rotation). The Oa and Ob helices move outwards by a 17° rotation.

  3. 3.

    The 30° rotation of the thumb domain in Pol ν

    The conformational changes between Ndna2 (closed finger and thumb-in) and Ndna4 (closed finger and thumb-out) structure. Y686, shown in stick-and-ball model, is a placeholder for the incoming templating base.

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DOI

https://doi.org/10.1038/nsmb.2985

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