Structure of the Full-length Human RPA14/32 Complex Gives Insights into the Mechanism of DNA Binding and Complex Formation

Xiaoyi Deng; Jeff E Habel; Venkataramen Kabaleeswaran; Edward H Snell; Marc S Wold; Gloria E.O Borgstahl

doi:10.1016/j.jmb.2007.09.074

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Structure of the Full-length Human RPA14/32 Complex Gives Insights into the Mechanism of DNA Binding and Complex Formation

Journal article

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Structure of the Full-length Human RPA14/32 Complex Gives Insights into the Mechanism of DNA Binding and Complex Formation

Xiaoyi Deng, Jeff E Habel, Venkataramen Kabaleeswaran, Edward H Snell, Marc S Wold and Gloria E.O Borgstahl

Journal of molecular biology, Vol.374(4), pp.865-876

12/07/2007

DOI: 10.1016/j.jmb.2007.09.074

PMID: 17976647

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Abstract

Replication protein A (RPA) is the ubiquitous, eukaryotic single-stranded DNA (ssDNA) binding protein and is essential for DNA replication, recombination, and repair. Here, crystal structures of the soluble RPA heterodimer, composed of the RPA14 and RPA32 subunits, have been determined for the full-length protein in multiple crystal forms. In all crystals, the electron density for the N-terminal (residues 1–42) and C-terminal (residues 175–270) regions of RPA32 is weak and of poor quality indicating that these regions are disordered and/or assume multiple positions in the crystals. Hence, the RPA32 N terminus, that is hyperphosphorylated in a cell-cycle-dependent manner and in response to DNA damaging agents, appears to be inherently disordered in the unphosphorylated state. The C-terminal, winged helix-loop-helix, protein–protein interaction domain adopts several conformations perhaps to facilitate its interaction with various proteins. Although the ordered regions of RPA14/32 resemble the previously solved protease-resistant core crystal structure, the quaternary structures between the heterodimers are quite different. Thus, the four-helix bundle quaternary assembly noted in the original core structure is unlikely to be related to the quaternary structure of the intact heterotrimer. An organic ligand binding site between subunits RPA14 and RPA32 was identified to bind dioxane. Comparison of the ssDNA binding surfaces of RPA70 with RPA14/32 showed that the lower affinity of RPA14/32 can be attributed to a shallower binding crevice with reduced positive electrostatic charge.

X-ray crystallography

replication protein A

helix bundle

subunit interaction

OB-fold

Details

Title: Subtitle: Structure of the Full-length Human RPA14/32 Complex Gives Insights into the Mechanism of DNA Binding and Complex Formation
Creators: Xiaoyi Deng - The Eppley Institute for Research in Cancer and Allied Diseases, 987696 Nebraska Medical Center, Omaha, NE 68198-7696, USA
Jeff E Habel - The University of Toledo, Department of Chemistry, 2801 W. Bancroft Street, Toledo, OH 43606, USA
Venkataramen Kabaleeswaran - The University of Toledo, Department of Chemistry, 2801 W. Bancroft Street, Toledo, OH 43606, USA
Edward H Snell - Hauptman-Woodward Medical Research Institute, Buffalo, NY; Department of Structural Biology, SUNY at Buffalo, Buffalo, NY 14203, USA
Marc S Wold - Department of Biochemistry, University of Iowa College of Medicine, 3107 MERF, Iowa City Iowa 52242-2600,, USA
Gloria E.O Borgstahl - The Eppley Institute for Research in Cancer and Allied Diseases, 987696 Nebraska Medical Center, Omaha, NE 68198-7696, USA
Resource Type: Journal article
Publication Details: Journal of molecular biology, Vol.374(4), pp.865-876
Publisher: Elsevier Ltd
DOI: 10.1016/j.jmb.2007.09.074
PMID: 17976647
ISSN: 0022-2836
eISSN: 1089-8638
Language: English
Date published: 12/07/2007
Academic Unit: Radiation Oncology; Biochemistry and Molecular Biology
Record Identifier: 9984024409402771

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