A specific subdomain in 29 DNA polymerase confers both processivity and strand displacement capacity

Recent crystallographic studies of φ29 DNA polymerase have provided structural insights into its strand displacement and processivity. A specific insertion named terminal protein region 2 (TPR2), present only in protein-primed DNA polymerases, together with the exonuclease, thumb, and palm subdomain...

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Autores: Rodríguez García, Irene, Lázaro, José M., Blanco Dávila, Luis, Kamtekar, S., Berman, Andrea J., Wang, J., Steitz, T. A., Salas, Margarita, Vega, Miguel de
Tipo de recurso: artículo
Fecha de publicación:2005
País:España
Institución:Consejo Superior de Investigaciones Científicas (CSIC)
Repositorio:DIGITAL.CSIC. Repositorio Institucional del CSIC
OAI Identifier:oai:digital.csic.es:10261/37712
Acceso en línea:http://hdl.handle.net/10261/37712
Access Level:acceso abierto
Palabra clave:Protein-primed replication
Terminal protein region
Helicase-like activity
DNA-binding stability
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spelling A specific subdomain in 29 DNA polymerase confers both processivity and strand displacement capacityRodríguez García, IreneLázaro, José M.Blanco Dávila, LuisKamtekar, S.Berman, Andrea J.Wang, J.Steitz, T. A.Salas, MargaritaVega, Miguel deProtein-primed replicationTerminal protein regionHelicase-like activityDNA-binding stabilityRecent crystallographic studies of φ29 DNA polymerase have provided structural insights into its strand displacement and processivity. A specific insertion named terminal protein region 2 (TPR2), present only in protein-primed DNA polymerases, together with the exonuclease, thumb, and palm subdomains, forms two tori capable of interacting with DNA. To analyze the functional role of this insertion, we constructed a φ29 DNA polymerase deletion mutant lacking TPR2 amino acid residues Asp-398 to Glu-420. Biochemical analysis of the mutant DNA polymerase indicates that its DNA-binding capacity is diminished, drastically decreasing its processivity. In addition, removal of the TPR2 insertion abolishes the intrinsic capacity of φ29 DNA polymerase to perform strand displacement coupled to DNA synthesis. Therefore, the biochemical results described here directly demonstrate that TPR2 plays a critical role in strand displacement and processivity.This investigation was aided by Research Grant BMC 2002-03818 from the Spanish Ministry of Science and Technology (to M.S.), Grant R01GM57510 from the National Institutes of Health (to T.A.S.), and an institutional grant from Fundación Ramón Areces to the Centro de Biología Molecular “Severo Ochoa.” I.R. was a predoctoral fellow of the Consejo Superior de Investigaciones Científicas.Peer reviewedNational Academy of Sciences (U.S.)Ministerio de Ciencia y Tecnología (España)National Institutes of Health (US)Fundación Ramón ArecesConsejo Superior de Investigaciones Científicas (España)201120112005info:eu-repo/semantics/articlehttp://purl.org/coar/resource_type/c_6501http://hdl.handle.net/10261/37712reponame:DIGITAL.CSIC. Repositorio Institucional del CSICinstname:Consejo Superior de Investigaciones Científicas (CSIC)Ingléshttp://dx.doi.org/10.1073/pnas.0500597102info:eu-repo/semantics/openAccessoai:digital.csic.es:10261/377122026-05-22T06:33:51Z
dc.title.none.fl_str_mv A specific subdomain in 29 DNA polymerase confers both processivity and strand displacement capacity
title A specific subdomain in 29 DNA polymerase confers both processivity and strand displacement capacity
spellingShingle A specific subdomain in 29 DNA polymerase confers both processivity and strand displacement capacity
Rodríguez García, Irene
Protein-primed replication
Terminal protein region
Helicase-like activity
DNA-binding stability
title_short A specific subdomain in 29 DNA polymerase confers both processivity and strand displacement capacity
title_full A specific subdomain in 29 DNA polymerase confers both processivity and strand displacement capacity
title_fullStr A specific subdomain in 29 DNA polymerase confers both processivity and strand displacement capacity
title_full_unstemmed A specific subdomain in 29 DNA polymerase confers both processivity and strand displacement capacity
title_sort A specific subdomain in 29 DNA polymerase confers both processivity and strand displacement capacity
dc.creator.none.fl_str_mv Rodríguez García, Irene
Lázaro, José M.
Blanco Dávila, Luis
Kamtekar, S.
Berman, Andrea J.
Wang, J.
Steitz, T. A.
Salas, Margarita
Vega, Miguel de
author Rodríguez García, Irene
author_facet Rodríguez García, Irene
Lázaro, José M.
Blanco Dávila, Luis
Kamtekar, S.
Berman, Andrea J.
Wang, J.
Steitz, T. A.
Salas, Margarita
Vega, Miguel de
author_role author
author2 Lázaro, José M.
Blanco Dávila, Luis
Kamtekar, S.
Berman, Andrea J.
Wang, J.
Steitz, T. A.
Salas, Margarita
Vega, Miguel de
author2_role author
author
author
author
author
author
author
author
dc.contributor.none.fl_str_mv Ministerio de Ciencia y Tecnología (España)
National Institutes of Health (US)
Fundación Ramón Areces
Consejo Superior de Investigaciones Científicas (España)
dc.subject.none.fl_str_mv Protein-primed replication
Terminal protein region
Helicase-like activity
DNA-binding stability
topic Protein-primed replication
Terminal protein region
Helicase-like activity
DNA-binding stability
description Recent crystallographic studies of φ29 DNA polymerase have provided structural insights into its strand displacement and processivity. A specific insertion named terminal protein region 2 (TPR2), present only in protein-primed DNA polymerases, together with the exonuclease, thumb, and palm subdomains, forms two tori capable of interacting with DNA. To analyze the functional role of this insertion, we constructed a φ29 DNA polymerase deletion mutant lacking TPR2 amino acid residues Asp-398 to Glu-420. Biochemical analysis of the mutant DNA polymerase indicates that its DNA-binding capacity is diminished, drastically decreasing its processivity. In addition, removal of the TPR2 insertion abolishes the intrinsic capacity of φ29 DNA polymerase to perform strand displacement coupled to DNA synthesis. Therefore, the biochemical results described here directly demonstrate that TPR2 plays a critical role in strand displacement and processivity.
publishDate 2005
dc.date.none.fl_str_mv 2005
2011
2011
dc.type.none.fl_str_mv info:eu-repo/semantics/article
http://purl.org/coar/resource_type/c_6501
format article
dc.identifier.none.fl_str_mv http://hdl.handle.net/10261/37712
url http://hdl.handle.net/10261/37712
dc.language.none.fl_str_mv Inglés
language_invalid_str_mv Inglés
dc.relation.none.fl_str_mv http://dx.doi.org/10.1073/pnas.0500597102
dc.rights.none.fl_str_mv info:eu-repo/semantics/openAccess
eu_rights_str_mv openAccess
dc.publisher.none.fl_str_mv National Academy of Sciences (U.S.)
publisher.none.fl_str_mv National Academy of Sciences (U.S.)
dc.source.none.fl_str_mv reponame:DIGITAL.CSIC. Repositorio Institucional del CSIC
instname:Consejo Superior de Investigaciones Científicas (CSIC)
instname_str Consejo Superior de Investigaciones Científicas (CSIC)
reponame_str DIGITAL.CSIC. Repositorio Institucional del CSIC
collection DIGITAL.CSIC. Repositorio Institucional del CSIC
repository.name.fl_str_mv
repository.mail.fl_str_mv
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score 15,811543

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