Cargando…

Solution-state structure of a fully alternately 2′-F/2′-OMe modified 42-nt dimeric siRNA construct

A high-resolution solution structure of a stable 42-nt RNA dimeric construct has been derived based on a high number of NMR observables including nuclear overhauser effects (NOEs), J-coupling constants and residual dipolar couplings (RDCs), which were all obtained with isotopically unlabeled molecul...

Descripción completa

Detalles Bibliográficos
Autores principales: Podbevsek, Peter, Allerson, Charles R., Bhat, Balkrishen, Plavec, Janez
Formato: Texto
Lenguaje:English
Publicado: Oxford University Press 2010
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC2978378/
https://www.ncbi.nlm.nih.gov/pubmed/20624819
http://dx.doi.org/10.1093/nar/gkq621
_version_ 1782191253951086592
author Podbevsek, Peter
Allerson, Charles R.
Bhat, Balkrishen
Plavec, Janez
author_facet Podbevsek, Peter
Allerson, Charles R.
Bhat, Balkrishen
Plavec, Janez
author_sort Podbevsek, Peter
collection PubMed
description A high-resolution solution structure of a stable 42-nt RNA dimeric construct has been derived based on a high number of NMR observables including nuclear overhauser effects (NOEs), J-coupling constants and residual dipolar couplings (RDCs), which were all obtained with isotopically unlabeled molecules. Two 21-nt siRNA that efficiently hybridize consist of ribose units that were alternately substituted by 2′-fluoro or 2′-methoxy groups. Structure calculations utilized a set of H-F RDC values for all 21 2′-fluoro modified nucleotides under conditions of weak alignment achieved by Pf1 phages. A completely 2′-F/2′-OMe modified dimeric RNA construct adopts an antiparallel double-helical structure consisting of 19 Watson–Crick base pairs with additional 3′ UU overhangs and a 5′ phosphate group on the antisense strand. NMR data suggest that the stability of individual base pairs is not uniform throughout the construct. While most of the double helical segment exhibits well dispersed imino resonances, the last three base pairs either display uncharacteristic chemical shifts of imino protons or absence of imino resonances even at lower temperatures. Accessibility of imino protons to solvent exchange suggests a difference in stability of duplex ends, which might be of importance for incorporation of the guide siRNA strand into a RISC.
format Text
id pubmed-2978378
institution National Center for Biotechnology Information
language English
publishDate 2010
publisher Oxford University Press
record_format MEDLINE/PubMed
spelling pubmed-29783782010-11-12 Solution-state structure of a fully alternately 2′-F/2′-OMe modified 42-nt dimeric siRNA construct Podbevsek, Peter Allerson, Charles R. Bhat, Balkrishen Plavec, Janez Nucleic Acids Res Structural Biology A high-resolution solution structure of a stable 42-nt RNA dimeric construct has been derived based on a high number of NMR observables including nuclear overhauser effects (NOEs), J-coupling constants and residual dipolar couplings (RDCs), which were all obtained with isotopically unlabeled molecules. Two 21-nt siRNA that efficiently hybridize consist of ribose units that were alternately substituted by 2′-fluoro or 2′-methoxy groups. Structure calculations utilized a set of H-F RDC values for all 21 2′-fluoro modified nucleotides under conditions of weak alignment achieved by Pf1 phages. A completely 2′-F/2′-OMe modified dimeric RNA construct adopts an antiparallel double-helical structure consisting of 19 Watson–Crick base pairs with additional 3′ UU overhangs and a 5′ phosphate group on the antisense strand. NMR data suggest that the stability of individual base pairs is not uniform throughout the construct. While most of the double helical segment exhibits well dispersed imino resonances, the last three base pairs either display uncharacteristic chemical shifts of imino protons or absence of imino resonances even at lower temperatures. Accessibility of imino protons to solvent exchange suggests a difference in stability of duplex ends, which might be of importance for incorporation of the guide siRNA strand into a RISC. Oxford University Press 2010-11 2010-07-12 /pmc/articles/PMC2978378/ /pubmed/20624819 http://dx.doi.org/10.1093/nar/gkq621 Text en © The Author(s) 2010. Published by Oxford University Press. http://creativecommons.org/licenses/by-nc/2.5 This is an Open Access article distributed under the terms of the Creative Commons Attribution Non-Commercial License (http://creativecommons.org/licenses/by-nc/2.5), which permits unrestricted non-commercial use, distribution, and reproduction in any medium, provided the original work is properly cited.
spellingShingle Structural Biology
Podbevsek, Peter
Allerson, Charles R.
Bhat, Balkrishen
Plavec, Janez
Solution-state structure of a fully alternately 2′-F/2′-OMe modified 42-nt dimeric siRNA construct
title Solution-state structure of a fully alternately 2′-F/2′-OMe modified 42-nt dimeric siRNA construct
title_full Solution-state structure of a fully alternately 2′-F/2′-OMe modified 42-nt dimeric siRNA construct
title_fullStr Solution-state structure of a fully alternately 2′-F/2′-OMe modified 42-nt dimeric siRNA construct
title_full_unstemmed Solution-state structure of a fully alternately 2′-F/2′-OMe modified 42-nt dimeric siRNA construct
title_short Solution-state structure of a fully alternately 2′-F/2′-OMe modified 42-nt dimeric siRNA construct
title_sort solution-state structure of a fully alternately 2′-f/2′-ome modified 42-nt dimeric sirna construct
topic Structural Biology
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC2978378/
https://www.ncbi.nlm.nih.gov/pubmed/20624819
http://dx.doi.org/10.1093/nar/gkq621
work_keys_str_mv AT podbevsekpeter solutionstatestructureofafullyalternately2f2omemodified42ntdimericsirnaconstruct
AT allersoncharlesr solutionstatestructureofafullyalternately2f2omemodified42ntdimericsirnaconstruct
AT bhatbalkrishen solutionstatestructureofafullyalternately2f2omemodified42ntdimericsirnaconstruct
AT plavecjanez solutionstatestructureofafullyalternately2f2omemodified42ntdimericsirnaconstruct