Structural Insights into Yeast DNA Polymerase δ by Small Angle X-ray Scattering

Rinku Jain; Michal Hammel; Robert Johnson; Louise Prakash; Satya Prakash; Aneel K. Aggarwal

doi:10.1016/j.jmb.2009.09.066

Structural Insights into Yeast DNA Polymerase δ by Small Angle X-ray Scattering

Rinku Jain, Michal Hammel, Robert Johnson, Louise Prakash, Satya Prakash, Aneel K. Aggarwal

Biochemistry & Molecular Biology

Research output: Contribution to journal › Article › peer-review

27 Scopus citations

Abstract

DNA polymerase δ (Polδ) is a multisubunit polymerase that plays an indispensable role in replication from yeast to humans. Polδ from Saccharomyces cerevisiae is composed of three subunits: Pol3, Pol31, and Pol32. Despite the elucidation of the structures and models of the individual subunits (or portions, thereof), the nature of their assembly remains unclear. We present here a small-angle X-ray scattering analysis of a yeast Polδ complex (Polδ_T) composed of Pol3, Pol31, and Pol32N (amino acids 1-103 of Pol32). From the small angle X-ray scattering global parameters and reconstructed envelopes, we show that Polδ_T adopts an elongated conformation with a radius of gyration (R_g) of ∼ 52 Å and a maximal dimension of ∼ 190 Å. We also propose an orientation for the accessory Pol31-Pol32N subunits relative to the Pol3 catalytic core that best agrees with the experimental scattering profile. The analysis also points to significant conformational variability that may allow Polδ to better coordinate its action with other proteins at the replication fork.

Original language	English (US)
Pages (from-to)	377-382
Number of pages	6
Journal	Journal of Molecular Biology
Volume	394
Issue number	3
DOIs	https://doi.org/10.1016/j.jmb.2009.09.066
State	Published - Dec 4 2009

Keywords

DNA polymerase
DNA replication
cancer
mutagenesis

ASJC Scopus subject areas

Molecular Biology
Biophysics
Structural Biology

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10.1016/j.jmb.2009.09.066

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title = "Structural Insights into Yeast DNA Polymerase δ by Small Angle X-ray Scattering",

abstract = "DNA polymerase δ (Polδ) is a multisubunit polymerase that plays an indispensable role in replication from yeast to humans. Polδ from Saccharomyces cerevisiae is composed of three subunits: Pol3, Pol31, and Pol32. Despite the elucidation of the structures and models of the individual subunits (or portions, thereof), the nature of their assembly remains unclear. We present here a small-angle X-ray scattering analysis of a yeast Polδ complex (PolδT) composed of Pol3, Pol31, and Pol32N (amino acids 1-103 of Pol32). From the small angle X-ray scattering global parameters and reconstructed envelopes, we show that PolδT adopts an elongated conformation with a radius of gyration (Rg) of ∼ 52 {\AA} and a maximal dimension of ∼ 190 {\AA}. We also propose an orientation for the accessory Pol31-Pol32N subunits relative to the Pol3 catalytic core that best agrees with the experimental scattering profile. The analysis also points to significant conformational variability that may allow Polδ to better coordinate its action with other proteins at the replication fork.",

keywords = "DNA polymerase, DNA replication, cancer, mutagenesis",

author = "Rinku Jain and Michal Hammel and Robert Johnson and Louise Prakash and Satya Prakash and Aggarwal, {Aneel K.}",

note = "Funding Information: We thank the Berkeley Lab Advanced Light Source and SIBYLS staff at beamline 12.3.1 for provision of X-ray facilities, and S. Chakravarty for help with modeling. This work was supported, in part, by grants from the Office of Science, Ofﬁce of Biological and Environmental Research, US Department of Energy , under contract DE-AC02-05CH11231 for SIBLYS beamline efforts, and by grant CA138546 from the US National Institutes of Health . The computational part of SAXS analysis was supported by Laboratory Research Computing at Lawrence Berkeley National Laboratory (Berkeley, CA).",

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AU - Jain, Rinku

AU - Hammel, Michal

AU - Johnson, Robert

AU - Prakash, Louise

AU - Prakash, Satya

AU - Aggarwal, Aneel K.

N1 - Funding Information: We thank the Berkeley Lab Advanced Light Source and SIBYLS staff at beamline 12.3.1 for provision of X-ray facilities, and S. Chakravarty for help with modeling. This work was supported, in part, by grants from the Office of Science, Ofﬁce of Biological and Environmental Research, US Department of Energy , under contract DE-AC02-05CH11231 for SIBLYS beamline efforts, and by grant CA138546 from the US National Institutes of Health . The computational part of SAXS analysis was supported by Laboratory Research Computing at Lawrence Berkeley National Laboratory (Berkeley, CA).

PY - 2009/12/4

Y1 - 2009/12/4

N2 - DNA polymerase δ (Polδ) is a multisubunit polymerase that plays an indispensable role in replication from yeast to humans. Polδ from Saccharomyces cerevisiae is composed of three subunits: Pol3, Pol31, and Pol32. Despite the elucidation of the structures and models of the individual subunits (or portions, thereof), the nature of their assembly remains unclear. We present here a small-angle X-ray scattering analysis of a yeast Polδ complex (PolδT) composed of Pol3, Pol31, and Pol32N (amino acids 1-103 of Pol32). From the small angle X-ray scattering global parameters and reconstructed envelopes, we show that PolδT adopts an elongated conformation with a radius of gyration (Rg) of ∼ 52 Å and a maximal dimension of ∼ 190 Å. We also propose an orientation for the accessory Pol31-Pol32N subunits relative to the Pol3 catalytic core that best agrees with the experimental scattering profile. The analysis also points to significant conformational variability that may allow Polδ to better coordinate its action with other proteins at the replication fork.

AB - DNA polymerase δ (Polδ) is a multisubunit polymerase that plays an indispensable role in replication from yeast to humans. Polδ from Saccharomyces cerevisiae is composed of three subunits: Pol3, Pol31, and Pol32. Despite the elucidation of the structures and models of the individual subunits (or portions, thereof), the nature of their assembly remains unclear. We present here a small-angle X-ray scattering analysis of a yeast Polδ complex (PolδT) composed of Pol3, Pol31, and Pol32N (amino acids 1-103 of Pol32). From the small angle X-ray scattering global parameters and reconstructed envelopes, we show that PolδT adopts an elongated conformation with a radius of gyration (Rg) of ∼ 52 Å and a maximal dimension of ∼ 190 Å. We also propose an orientation for the accessory Pol31-Pol32N subunits relative to the Pol3 catalytic core that best agrees with the experimental scattering profile. The analysis also points to significant conformational variability that may allow Polδ to better coordinate its action with other proteins at the replication fork.

KW - DNA polymerase

KW - DNA replication

KW - cancer

KW - mutagenesis

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Structural Insights into Yeast DNA Polymerase δ by Small Angle X-ray Scattering

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