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Nat Commun. 2017 May 2;8:15075. doi: 10.1038/ncomms15075.

Identification and characterization of a heterotrimeric archaeal DNA polymerase holoenzyme.

Author information

1
Department of Molecular and Cellular Biochemistry, Indiana University, Simon Hall MSB, 212 S Hawthorne Dr, Bloomington, Indiana 47405, USA.
2
Sir William Dunn School of Pathology, University of Oxford, South Parks Road, Oxford OX1 3RE, UK.
3
Structural Biology Unit, CIC bioGUNE, CIBERehd, 48160 Derio, Spain.
4
New England Biolabs, 240 County Road, Ipswich, Massachusetts 01938-2723, USA.
5
Department of Chemistry, Indiana University, Bloomington, Indiana 47405, USA.
6
Molecular Biology of Archaea, Institute of Biology II, Microbiology, University of Freiburg, Schaenzlestr. 1, 79104 Freiburg, Germany.
7
Department of Biochemistry, Oxford Glycobiology Institute, University of Oxford, South Parks Road, Oxford OX1 3QU, UK.
8
IKERBASQUE, Basque Foundation for Science, 48013 Bilbao, Spain.
9
Department of Biology, Indiana University, Simon Hall MSB, 212 S Hawthorne Dr, Bloomington, Indiana 47405, USA.

Abstract

Since their initial characterization over 30 years ago, it has been believed that the archaeal B-family DNA polymerases are single-subunit enzymes. This contrasts with the multi-subunit B-family replicative polymerases of eukaryotes. Here we reveal that the highly studied PolB1 from Sulfolobus solfataricus exists as a heterotrimeric complex in cell extracts. Two small subunits, PBP1 and PBP2, associate with distinct surfaces of the larger catalytic subunit and influence the enzymatic properties of the DNA polymerase. Thus, multi-subunit replicative DNA polymerase holoenzymes are present in all three domains of life. We reveal the architecture of the assembly by a combination of cross-linking coupled with mass spectrometry, X-ray crystallography and single-particle electron microscopy. The small subunits stabilize the holoenzyme assembly and the acidic tail of one small subunit mitigates the ability of the enzyme to perform strand-displacement synthesis, with important implications for lagging strand DNA synthesis.

PMID:
28462924
PMCID:
PMC5418573
DOI:
10.1038/ncomms15075
[Indexed for MEDLINE]
Free PMC Article

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