2'-Alkynyl spin-labelling is a minimally perturbing tool for DNA structural analysis

Jack Hardwick; Marius Haugland; Afaf El-Sagheer; Denis Ptchelkine; Frank Beierlein; Andrew Lane; Tom Brown; Janet E. Lovett; Edward Anderson

doi:10.1093/nar/gkaa086

2'-Alkynyl spin-labelling is a minimally perturbing tool for DNA structural analysis

Jack Hardwick, Marius Haugland, Afaf El-Sagheer, Denis Ptchelkine, Frank Beierlein, Andrew Lane, Tom Brown, Janet E. Lovett, Edward Anderson^*

^*Corresponding author for this work

Research output: Contribution to journal › Article › peer-review

Abstract

The determination of distances between specific points in nucleic acids is essential to understanding their behaviour at the molecular level. The ability to measure distances of 2–10 nm is particularly important: deformations arising from protein binding commonly fall within this range, but the reliable measurement of such distances for a conformational ensemble remains a significant challenge. Using several techniques, we show that electron paramagnetic resonance (EPR) spectroscopy of oligonucleotides spin-labelled with triazole-appended nitroxides at the 2′ position offers a robust and minimally perturbing tool for obtaining such measurements. For two nitroxides, we present results from EPR spectroscopy, X-ray crystal structures of B-form spin-labelled DNA duplexes, molecular dynamics simulations and nuclear magnetic resonance spectroscopy. These four methods are mutually supportive, and pinpoint the locations of the spin labels on the duplexes. In doing so, this work establishes 2′-alkynyl nitroxide spin-labelling as a minimally perturbing method for probing DNA conformation.

Original language	English
Article number	gkaa086
Journal	Nucleic Acids Research
Volume	Advance articles
Early online date	13 Feb 2020
DOIs	https://doi.org/10.1093/nar/gkaa086
Publication status	E-pub ahead of print - 13 Feb 2020

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10.1093/nar/gkaa086Licence: CC BY

Hardwick_2020_NAR_2'-alkynyl_CC
Copyright © The Author(s) 2020. Published by Oxford University Press on behalf of Nucleic Acids Research. This is an Open Access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/4.0/), which permits unrestricted reuse, distribution, and reproduction in any medium, provided the original work is properly cited.
Final published version, 3.62 MBLicence: CC BY

Cite this

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title = "2'-Alkynyl spin-labelling is a minimally perturbing tool for DNA structural analysis",

abstract = "The determination of distances between specific points in nucleic acids is essential to understanding their behaviour at the molecular level. The ability to measure distances of 2–10 nm is particularly important: deformations arising from protein binding commonly fall within this range, but the reliable measurement of such distances for a conformational ensemble remains a significant challenge. Using several techniques, we show that electron paramagnetic resonance (EPR) spectroscopy of oligonucleotides spin-labelled with triazole-appended nitroxides at the 2′ position offers a robust and minimally perturbing tool for obtaining such measurements. For two nitroxides, we present results from EPR spectroscopy, X-ray crystal structures of B-form spin-labelled DNA duplexes, molecular dynamics simulations and nuclear magnetic resonance spectroscopy. These four methods are mutually supportive, and pinpoint the locations of the spin labels on the duplexes. In doing so, this work establishes 2′-alkynyl nitroxide spin-labelling as a minimally perturbing method for probing DNA conformation.",

author = "Jack Hardwick and Marius Haugland and Afaf El-Sagheer and Denis Ptchelkine and Frank Beierlein and Andrew Lane and Tom Brown and Lovett, \{Janet E.\} and Edward Anderson",

note = "Funding: Engineering and Physical Sciences Research Council [EP/M019195/1]; Engineering and Physical Sciences Research Council Studentship (to J.S.H.); Biotechnology and Biological Sciences Research Council [BB/J001694/2, BB/R021848/1]; ADTBio; University of Kentucky and NCI Cancer Center Support Grant [P30 CA177558]; The Carmen L. Buck Endowment; Emerging Fields Initiative of the Friedrich-Alexander-University of Erlangen-Nuremberg [Grant title {\textquoteleft}Chemistry in Live Cells{\textquoteright}]; Wellcome Trust [099149/Z/12/Z]; Royal Society, University Research Fellowship (to J.E.L.). Funding for open access charge: University of Oxford.",

year = "2020",

month = feb,

day = "13",

doi = "10.1093/nar/gkaa086",

language = "English",

volume = "Advance articles",

journal = "Nucleic Acids Research",

issn = "0305-1048",

publisher = "Oxford University Press",

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TY - JOUR

T1 - 2'-Alkynyl spin-labelling is a minimally perturbing tool for DNA structural analysis

AU - Hardwick, Jack

AU - Haugland, Marius

AU - El-Sagheer, Afaf

AU - Ptchelkine, Denis

AU - Beierlein, Frank

AU - Lane, Andrew

AU - Brown, Tom

AU - Lovett, Janet E.

AU - Anderson, Edward

N1 - Funding: Engineering and Physical Sciences Research Council [EP/M019195/1]; Engineering and Physical Sciences Research Council Studentship (to J.S.H.); Biotechnology and Biological Sciences Research Council [BB/J001694/2, BB/R021848/1]; ADTBio; University of Kentucky and NCI Cancer Center Support Grant [P30 CA177558]; The Carmen L. Buck Endowment; Emerging Fields Initiative of the Friedrich-Alexander-University of Erlangen-Nuremberg [Grant title ‘Chemistry in Live Cells’]; Wellcome Trust [099149/Z/12/Z]; Royal Society, University Research Fellowship (to J.E.L.). Funding for open access charge: University of Oxford.

PY - 2020/2/13

Y1 - 2020/2/13

N2 - The determination of distances between specific points in nucleic acids is essential to understanding their behaviour at the molecular level. The ability to measure distances of 2–10 nm is particularly important: deformations arising from protein binding commonly fall within this range, but the reliable measurement of such distances for a conformational ensemble remains a significant challenge. Using several techniques, we show that electron paramagnetic resonance (EPR) spectroscopy of oligonucleotides spin-labelled with triazole-appended nitroxides at the 2′ position offers a robust and minimally perturbing tool for obtaining such measurements. For two nitroxides, we present results from EPR spectroscopy, X-ray crystal structures of B-form spin-labelled DNA duplexes, molecular dynamics simulations and nuclear magnetic resonance spectroscopy. These four methods are mutually supportive, and pinpoint the locations of the spin labels on the duplexes. In doing so, this work establishes 2′-alkynyl nitroxide spin-labelling as a minimally perturbing method for probing DNA conformation.

AB - The determination of distances between specific points in nucleic acids is essential to understanding their behaviour at the molecular level. The ability to measure distances of 2–10 nm is particularly important: deformations arising from protein binding commonly fall within this range, but the reliable measurement of such distances for a conformational ensemble remains a significant challenge. Using several techniques, we show that electron paramagnetic resonance (EPR) spectroscopy of oligonucleotides spin-labelled with triazole-appended nitroxides at the 2′ position offers a robust and minimally perturbing tool for obtaining such measurements. For two nitroxides, we present results from EPR spectroscopy, X-ray crystal structures of B-form spin-labelled DNA duplexes, molecular dynamics simulations and nuclear magnetic resonance spectroscopy. These four methods are mutually supportive, and pinpoint the locations of the spin labels on the duplexes. In doing so, this work establishes 2′-alkynyl nitroxide spin-labelling as a minimally perturbing method for probing DNA conformation.

UR - https://www.scopus.com/pages/publications/85082560953

U2 - 10.1093/nar/gkaa086

DO - 10.1093/nar/gkaa086

M3 - Article

SN - 0305-1048

VL - Advance articles

JO - Nucleic Acids Research

JF - Nucleic Acids Research

M1 - gkaa086

ER -

2'-Alkynyl spin-labelling is a minimally perturbing tool for DNA structural analysis

Abstract

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