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Title: Structural insights into the adaptation of proliferating cell nuclear antigen (PCNA) from Haloferax volcanii to a high-salt environment

Abstract

The crystal structure of PCNA from the halophilic archaeon H. volcanii reveals specific features of the charge distribution on the protein surface that reflect adaptation to a high-salt environment and suggests a different type of interaction with DNA in halophilic PCNAs. The sliding clamp proliferating cell nuclear antigen (PCNA) plays vital roles in many aspects of DNA replication and repair in eukaryotic cells and in archaea. Realising the full potential of archaea as a model for PCNA function requires a combination of biochemical and genetic approaches. In order to provide a platform for subsequent reverse genetic analysis, PCNA from the halophilic archaeon Haloferax volcanii was subjected to crystallographic analysis. The gene was cloned and expressed in Escherichia coli and the protein was purified by affinity chromatography and crystallized by the vapour-diffusion technique. The structure was determined by molecular replacement and refined at 3.5 Å resolution to a final R factor of 23.7% (R{sub free} = 25%). PCNA from H. volcanii was found to be homotrimeric and to resemble other homotrimeric PCNA clamps but with several differences that appear to be associated with adaptation of the protein to the high intracellular salt concentrations found in H. volcanii cells.

Authors:
 [1];  [2];  [2];  [1]
  1. Karolinska Institutet, NOVUM, Centre of Structural Biochemistry, S-14157 Huddinge (Sweden)
  2. Department of Biology, University of Copenhagen, Ole Maaløes Vej 5, 2200 Copenhagen (Denmark)
Publication Date:
OSTI Identifier:
22351195
Resource Type:
Journal Article
Journal Name:
Acta Crystallographica. Section D: Biological Crystallography
Additional Journal Information:
Journal Volume: 65; Journal Issue: Pt 10; Other Information: PMCID: PMC2756170; PMID: 19770505; PUBLISHER-ID: ea5108; OAI: oai:pubmedcentral.nih.gov:2756170; Copyright (c) International Union of Crystallography 2009; Country of input: International Atomic Energy Agency (IAEA); Journal ID: ISSN 0907-4449
Country of Publication:
Denmark
Language:
English
Subject:
75 CONDENSED MATTER PHYSICS, SUPERCONDUCTIVITY AND SUPERFLUIDITY; AFFINITY; CHARGE DISTRIBUTION; CRYSTAL STRUCTURE; DIFFUSION; DISTRIBUTION; ENVIRONMENT; ESCHERICHIA COLI; INTERACTIONS; IRON; POTENTIALS; PROTEINS; R FACTORS; RESOLUTION; SURFACES

Citation Formats

Morgunova, Ekaterina, Gray, Fiona C., MacNeill, Stuart A., Centre for Biomolecular Sciences, University of St Andrews, North Haugh, St Andrews, Fife KY16 9ST,Scotland, and Ladenstein, Rudolf. Structural insights into the adaptation of proliferating cell nuclear antigen (PCNA) from Haloferax volcanii to a high-salt environment. Denmark: N. p., 2009. Web. doi:10.1107/S0907444909029321.
Morgunova, Ekaterina, Gray, Fiona C., MacNeill, Stuart A., Centre for Biomolecular Sciences, University of St Andrews, North Haugh, St Andrews, Fife KY16 9ST,Scotland, & Ladenstein, Rudolf. Structural insights into the adaptation of proliferating cell nuclear antigen (PCNA) from Haloferax volcanii to a high-salt environment. Denmark. doi:10.1107/S0907444909029321.
Morgunova, Ekaterina, Gray, Fiona C., MacNeill, Stuart A., Centre for Biomolecular Sciences, University of St Andrews, North Haugh, St Andrews, Fife KY16 9ST,Scotland, and Ladenstein, Rudolf. Thu . "Structural insights into the adaptation of proliferating cell nuclear antigen (PCNA) from Haloferax volcanii to a high-salt environment". Denmark. doi:10.1107/S0907444909029321.
@article{osti_22351195,
title = {Structural insights into the adaptation of proliferating cell nuclear antigen (PCNA) from Haloferax volcanii to a high-salt environment},
author = {Morgunova, Ekaterina and Gray, Fiona C. and MacNeill, Stuart A. and Centre for Biomolecular Sciences, University of St Andrews, North Haugh, St Andrews, Fife KY16 9ST,Scotland and Ladenstein, Rudolf},
abstractNote = {The crystal structure of PCNA from the halophilic archaeon H. volcanii reveals specific features of the charge distribution on the protein surface that reflect adaptation to a high-salt environment and suggests a different type of interaction with DNA in halophilic PCNAs. The sliding clamp proliferating cell nuclear antigen (PCNA) plays vital roles in many aspects of DNA replication and repair in eukaryotic cells and in archaea. Realising the full potential of archaea as a model for PCNA function requires a combination of biochemical and genetic approaches. In order to provide a platform for subsequent reverse genetic analysis, PCNA from the halophilic archaeon Haloferax volcanii was subjected to crystallographic analysis. The gene was cloned and expressed in Escherichia coli and the protein was purified by affinity chromatography and crystallized by the vapour-diffusion technique. The structure was determined by molecular replacement and refined at 3.5 Å resolution to a final R factor of 23.7% (R{sub free} = 25%). PCNA from H. volcanii was found to be homotrimeric and to resemble other homotrimeric PCNA clamps but with several differences that appear to be associated with adaptation of the protein to the high intracellular salt concentrations found in H. volcanii cells.},
doi = {10.1107/S0907444909029321},
journal = {Acta Crystallographica. Section D: Biological Crystallography},
issn = {0907-4449},
number = Pt 10,
volume = 65,
place = {Denmark},
year = {2009},
month = {10}
}