Structure of human Rad51 protein filament from molecular modeling and site-specific linear dichroism spectroscopy

Handle URI:
http://hdl.handle.net/10754/599767
Title:
Structure of human Rad51 protein filament from molecular modeling and site-specific linear dichroism spectroscopy
Authors:
Reymer, A.; Frykholm, K.; Morimatsu, K.; Takahashi, M.; Norden, B.
Abstract:
To get mechanistic insight into the DNA strand-exchange reaction of homologous recombination, we solved a filament structure of a human Rad51 protein, combining molecular modeling with experimental data. We build our structure on reported structures for central and N-terminal parts of pure (uncomplexed) Rad51 protein by aid of linear dichroism spectroscopy, providing angular orientations of substituted tyrosine residues of Rad51-dsDNA filaments in solution. The structure, validated by comparison with an electron microscopy density map and results from mutation analysis, is proposed to represent an active solution structure of the nucleo-protein complex. An inhomogeneously stretched double-stranded DNA fitted into the filament emphasizes the strategic positioning of 2 putative DNA-binding loops in a way that allows us speculate about their possibly distinct roles in nucleo-protein filament assembly and DNA strand-exchange reaction. The model suggests that the extension of a single-stranded DNA molecule upon binding of Rad51 is ensured by intercalation of Tyr-232 of the L1 loop, which might act as a docking tool, aligning protein monomers along the DNA strand upon filament assembly. Arg-235, also sitting on L1, is in the right position to make electrostatic contact with the phosphate backbone of the other DNA strand. The L2 loop position and its more ordered compact conformation makes us propose that this loop has another role, as a binding site for an incoming double-stranded DNA. Our filament structure and spectroscopic approach open the possibility of analyzing details along the multistep path of the strand-exchange reaction.
Citation:
Reymer A, Frykholm K, Morimatsu K, Takahashi M, Norden B (2009) Structure of human Rad51 protein filament from molecular modeling and site-specific linear dichroism spectroscopy. Proceedings of the National Academy of Sciences 106: 13248–13253. Available: http://dx.doi.org/10.1073/pnas.0902723106.
Publisher:
Proceedings of the National Academy of Sciences
Journal:
Proceedings of the National Academy of Sciences
KAUST Grant Number:
KUK-11-008-23
Issue Date:
8-Jul-2009
DOI:
10.1073/pnas.0902723106
PubMed ID:
19587234
PubMed Central ID:
PMC2726390
Type:
Article
ISSN:
0027-8424; 1091-6490
Sponsors:
We thank Professor Edward H. Egelman (University of Virginia, Charlottesville, VA) for kindly providing us with the surface map of a 3D reconstruction of a HsRad51 filament electron micrograph and Dr. Axelle Renodon-Cornière for performing the recombinase activity measurements. This work was supported by King Abdullah University of Science and Technology Grant KUK-11-008-23 and the Association pour la Recherche sur le Cancer (M.T.).
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Full metadata record

DC FieldValue Language
dc.contributor.authorReymer, A.en
dc.contributor.authorFrykholm, K.en
dc.contributor.authorMorimatsu, K.en
dc.contributor.authorTakahashi, M.en
dc.contributor.authorNorden, B.en
dc.date.accessioned2016-02-28T06:09:20Zen
dc.date.available2016-02-28T06:09:20Zen
dc.date.issued2009-07-08en
dc.identifier.citationReymer A, Frykholm K, Morimatsu K, Takahashi M, Norden B (2009) Structure of human Rad51 protein filament from molecular modeling and site-specific linear dichroism spectroscopy. Proceedings of the National Academy of Sciences 106: 13248–13253. Available: http://dx.doi.org/10.1073/pnas.0902723106.en
dc.identifier.issn0027-8424en
dc.identifier.issn1091-6490en
dc.identifier.pmid19587234en
dc.identifier.doi10.1073/pnas.0902723106en
dc.identifier.urihttp://hdl.handle.net/10754/599767en
dc.description.abstractTo get mechanistic insight into the DNA strand-exchange reaction of homologous recombination, we solved a filament structure of a human Rad51 protein, combining molecular modeling with experimental data. We build our structure on reported structures for central and N-terminal parts of pure (uncomplexed) Rad51 protein by aid of linear dichroism spectroscopy, providing angular orientations of substituted tyrosine residues of Rad51-dsDNA filaments in solution. The structure, validated by comparison with an electron microscopy density map and results from mutation analysis, is proposed to represent an active solution structure of the nucleo-protein complex. An inhomogeneously stretched double-stranded DNA fitted into the filament emphasizes the strategic positioning of 2 putative DNA-binding loops in a way that allows us speculate about their possibly distinct roles in nucleo-protein filament assembly and DNA strand-exchange reaction. The model suggests that the extension of a single-stranded DNA molecule upon binding of Rad51 is ensured by intercalation of Tyr-232 of the L1 loop, which might act as a docking tool, aligning protein monomers along the DNA strand upon filament assembly. Arg-235, also sitting on L1, is in the right position to make electrostatic contact with the phosphate backbone of the other DNA strand. The L2 loop position and its more ordered compact conformation makes us propose that this loop has another role, as a binding site for an incoming double-stranded DNA. Our filament structure and spectroscopic approach open the possibility of analyzing details along the multistep path of the strand-exchange reaction.en
dc.description.sponsorshipWe thank Professor Edward H. Egelman (University of Virginia, Charlottesville, VA) for kindly providing us with the surface map of a 3D reconstruction of a HsRad51 filament electron micrograph and Dr. Axelle Renodon-Cornière for performing the recombinase activity measurements. This work was supported by King Abdullah University of Science and Technology Grant KUK-11-008-23 and the Association pour la Recherche sur le Cancer (M.T.).en
dc.publisherProceedings of the National Academy of Sciencesen
dc.subjectHomologous recombinationen
dc.subjectHsRad51en
dc.subject.meshModels, Molecularen
dc.titleStructure of human Rad51 protein filament from molecular modeling and site-specific linear dichroism spectroscopyen
dc.typeArticleen
dc.identifier.journalProceedings of the National Academy of Sciencesen
dc.identifier.pmcidPMC2726390en
dc.contributor.institutionChalmers University of Technology, Göteborg, Swedenen
dc.contributor.institutionUC Davis, Davis, United Statesen
dc.contributor.institutionUniversite de Nantes, Nantes, Franceen
kaust.grant.numberKUK-11-008-23en

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