Four ribose 2'-hydroxyl groups essential for catalytic function of the hairpin ribozyme

5 p.-4 fig.-1 tab.

Detalles Bibliográficos
Autores: Chowrira, Bharat M., Berzal-Herranz, Alfredo, Keller, Charles F., Burke, John M.
Tipo de recurso: artículo
Estado:Versión publicada
Fecha de publicación:1993
País:España
Institución:Consejo Superior de Investigaciones Científicas (CSIC)
Repositorio:DIGITAL.CSIC. Repositorio Institucional del CSIC
OAI Identifier:oai:digital.csic.es:10261/176317
Acceso en línea:http://hdl.handle.net/10261/176317
Access Level:acceso abierto
Palabra clave:Substrate binding
RNA cleavage
Tetrahymena
Sequence
DNA
Complementary
Resolution
Requirements
Ligation
Domain
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oai_identifier_str oai:digital.csic.es:10261/176317
network_acronym_str ES
network_name_str España
repository_id_str
spelling Four ribose 2'-hydroxyl groups essential for catalytic function of the hairpin ribozymeChowrira, Bharat M.Berzal-Herranz, AlfredoKeller, Charles F.Burke, John M.Substrate bindingRNA cleavageTetrahymenaSequenceDNAComplementaryResolutionRequirementsLigationDomain5 p.-4 fig.-1 tab.The hairpin ribozyme catalyzes site-specific cleavage of an RNA substrate using a magnesium-dependent transphosphorylation mechanism. Here, we describe experiments designed to test the importance of ribose 2'-hydroxyl groups for ribozyme function. Ribozymes for this work were synthesized in two segments using solid-phase RNA phosphoramidite chemistry. 2'-Deoxyribonucleotides were systematically introduced at each of the 50 positions within the ribozyme, and the catalytic activity of the resulting mixed RNA-DNA polymers was measured. Deletion of the 2'-hydroxyl group at each of four sites (A10, G11, A24, and C25) was found to result in severe inhibition of cleavage activity (kcat/KM decreased by 100- to 1000-fold), although KM measurements and mobility-shift assays showed that substrate binding was not affected. Identical results were obtained upon substitution of these ribonucleotides with 2'-O-methyl derivatives. Inhibition by 2'-modified sugars at G11 or A24 was rescued by increased Mg2+ concentrations, suggesting that these 2'-hydroxyls may function in magnesium binding. Our results demonstrate that the 2'-hydroxyl groups at A10, G11, A24, and C25 provide essential functions for catalysis, possibly forming important tertiary contacts or magnesium coordination sites that are necessary for active site architecture.This work was supported by grants from the National Institutes of HealthPeer reviewedAmerican Society for Biochemistry and Molecular BiologyNational Institutes of Health (US)Berzal-Herranz, Alfredo [0000-0003-3722-7973]Consejo Superior de Investigaciones Científicas [https://ror.org/02gfc7t72]201920191993info:eu-repo/semantics/articlehttp://purl.org/coar/resource_type/c_6501Publisher's versioninfo:eu-repo/semantics/publishedVersionhttp://hdl.handle.net/10261/176317reponame:DIGITAL.CSIC. Repositorio Institucional del CSICinstname:Consejo Superior de Investigaciones Científicas (CSIC)Ingléshttp://www.jbc.org/content/268/26/19458.longSíinfo:eu-repo/semantics/openAccessoai:digital.csic.es:10261/1763172026-05-22T06:33:51Z
dc.title.none.fl_str_mv Four ribose 2'-hydroxyl groups essential for catalytic function of the hairpin ribozyme
title Four ribose 2'-hydroxyl groups essential for catalytic function of the hairpin ribozyme
spellingShingle Four ribose 2'-hydroxyl groups essential for catalytic function of the hairpin ribozyme
Chowrira, Bharat M.
Substrate binding
RNA cleavage
Tetrahymena
Sequence
DNA
Complementary
Resolution
Requirements
Ligation
Domain
title_short Four ribose 2'-hydroxyl groups essential for catalytic function of the hairpin ribozyme
title_full Four ribose 2'-hydroxyl groups essential for catalytic function of the hairpin ribozyme
title_fullStr Four ribose 2'-hydroxyl groups essential for catalytic function of the hairpin ribozyme
title_full_unstemmed Four ribose 2'-hydroxyl groups essential for catalytic function of the hairpin ribozyme
title_sort Four ribose 2'-hydroxyl groups essential for catalytic function of the hairpin ribozyme
dc.creator.none.fl_str_mv Chowrira, Bharat M.
Berzal-Herranz, Alfredo
Keller, Charles F.
Burke, John M.
author Chowrira, Bharat M.
author_facet Chowrira, Bharat M.
Berzal-Herranz, Alfredo
Keller, Charles F.
Burke, John M.
author_role author
author2 Berzal-Herranz, Alfredo
Keller, Charles F.
Burke, John M.
author2_role author
author
author
dc.contributor.none.fl_str_mv National Institutes of Health (US)
Berzal-Herranz, Alfredo [0000-0003-3722-7973]
Consejo Superior de Investigaciones Científicas [https://ror.org/02gfc7t72]
dc.subject.none.fl_str_mv Substrate binding
RNA cleavage
Tetrahymena
Sequence
DNA
Complementary
Resolution
Requirements
Ligation
Domain
topic Substrate binding
RNA cleavage
Tetrahymena
Sequence
DNA
Complementary
Resolution
Requirements
Ligation
Domain
description 5 p.-4 fig.-1 tab.
publishDate 1993
dc.date.none.fl_str_mv 1993
2019
2019
dc.type.none.fl_str_mv info:eu-repo/semantics/article
http://purl.org/coar/resource_type/c_6501
Publisher's version
info:eu-repo/semantics/publishedVersion
format article
status_str publishedVersion
dc.identifier.none.fl_str_mv http://hdl.handle.net/10261/176317
url http://hdl.handle.net/10261/176317
dc.language.none.fl_str_mv Inglés
language_invalid_str_mv Inglés
dc.relation.none.fl_str_mv http://www.jbc.org/content/268/26/19458.long
Sí
dc.rights.none.fl_str_mv info:eu-repo/semantics/openAccess
eu_rights_str_mv openAccess
dc.publisher.none.fl_str_mv American Society for Biochemistry and Molecular Biology
publisher.none.fl_str_mv American Society for Biochemistry and Molecular Biology
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 15,812429

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