Protein-induced changes in DNA structure and dynamics observed with noncovalent site-directed spin-labeling and PELDOR

Gunnar Widtfeldt Reginsson; Sandip Shelke; Christophe Rouillon; Malcolm F White; Snorri T Sigurdsson; Olav Schiemann

doi:10.1093/nar/gks817

Protein-induced changes in DNA structure and dynamics observed with noncovalent site-directed spin-labeling and PELDOR

Gunnar Widtfeldt Reginsson, Sandip Shelke, Christophe Rouillon, Malcolm F White, Snorri T Sigurdsson, Olav Schiemann^*

^*Corresponding author for this work

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

Abstract

Site-directed spin labeling and pulsed electron–electron double resonance (PELDOR or DEER) have previously been applied successfully to study the structure and dynamics of nucleic acids. Spin labeling nucleic acids at specific sites requires the covalent attachment of spin labels, which involves rather complicated and laborious chemical synthesis. Here, we use a noncovalent label strategy that bypasses the covalent labeling chemistry and show that the binding specificity and efficiency are large enough to enable PELDOR or DEER measurements in DNA duplexes and a DNA duplex bound to the Lac repressor protein. In addition, the rigidity of the label not only allows resolution of the structure and dynamics of oligonucleotides but also the determination of label orientation and protein-induced conformational changes. The results prove that this labeling strategy in combination with PELDOR has a great potential for studying both structure and dynamics of oligonucleotides and their complexes with various ligands.

Original language	English
Article number	e11
Number of pages	10
Journal	Nucleic Acids Research
Volume	41
Issue number	1
Early online date	31 Aug 2012
DOIs	https://doi.org/10.1093/nar/gks817
Publication status	Published - Jan 2013

Keywords

Binding
PULSED ELECTRON
Nucleic-acids
Conformation
Lactose operon
Distance measurements
Operator
EPR spectroscopy
Electron-paramagnetic-resonance
LAC repressor

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10.1093/nar/gks817

Schiemann2012nar gks817protein induced
© The Author(s) 2012. Published by Oxford University Press. This is an Open Access article distributed under the terms of the Creative Commons Attribution Non-Commercial License (http://creativecommons.org/licenses/by-nc/3.0), which permits unrestricted non-commercial use, distribution, and reproduction in any medium, provided the original work is properly cited.
Final published version, 1.65 MB

Multifunctional molecular machines: multifunctional molecular machines acting on DNA
White, M. (PI) & Schiemann, O. (CoI)
The Wellcome Trust
1/11/10 → 31/10/15
Project: Standard
ConformationalStates of MembranceProtein: Conformational states of membrane proteins: Technology development for bioscience
Naismith, J. (PI) & Liu, H. (CoI)
BBSRC
1/07/10 → 9/01/16
Project: Standard
ConformationalStates of MembraneProtein: Conformational states of membranes proteins: Technology development for bioscience
Schiemann, O. (PI) & Cottrell, G. A. (CoI)
BBSRC
1/07/10 → 9/01/16
Project: Standard

Cite this

@article{0bde46b308e9486ea927da04195b2ff9,

title = "Protein-induced changes in DNA structure and dynamics observed with noncovalent site-directed spin-labeling and PELDOR",

abstract = "Site-directed spin labeling and pulsed electron–electron double resonance (PELDOR or DEER) have previously been applied successfully to study the structure and dynamics of nucleic acids. Spin labeling nucleic acids at specific sites requires the covalent attachment of spin labels, which involves rather complicated and laborious chemical synthesis. Here, we use a noncovalent label strategy that bypasses the covalent labeling chemistry and show that the binding specificity and efficiency are large enough to enable PELDOR or DEER measurements in DNA duplexes and a DNA duplex bound to the Lac repressor protein. In addition, the rigidity of the label not only allows resolution of the structure and dynamics of oligonucleotides but also the determination of label orientation and protein-induced conformational changes. The results prove that this labeling strategy in combination with PELDOR has a great potential for studying both structure and dynamics of oligonucleotides and their complexes with various ligands.",

keywords = "Binding, PULSED ELECTRON, Nucleic-acids, Conformation, Lactose operon, Distance measurements, Operator, EPR spectroscopy, Electron-paramagnetic-resonance, LAC repressor",

author = "Reginsson, {Gunnar Widtfeldt} and Sandip Shelke and Christophe Rouillon and White, {Malcolm F} and Sigurdsson, {Snorri T} and Olav Schiemann",

year = "2013",

month = jan,

doi = "10.1093/nar/gks817",

language = "English",

volume = "41",

journal = "Nucleic Acids Research",

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AU - Reginsson, Gunnar Widtfeldt

AU - Shelke, Sandip

AU - Rouillon, Christophe

AU - White, Malcolm F

AU - Sigurdsson, Snorri T

AU - Schiemann, Olav

PY - 2013/1

Y1 - 2013/1

N2 - Site-directed spin labeling and pulsed electron–electron double resonance (PELDOR or DEER) have previously been applied successfully to study the structure and dynamics of nucleic acids. Spin labeling nucleic acids at specific sites requires the covalent attachment of spin labels, which involves rather complicated and laborious chemical synthesis. Here, we use a noncovalent label strategy that bypasses the covalent labeling chemistry and show that the binding specificity and efficiency are large enough to enable PELDOR or DEER measurements in DNA duplexes and a DNA duplex bound to the Lac repressor protein. In addition, the rigidity of the label not only allows resolution of the structure and dynamics of oligonucleotides but also the determination of label orientation and protein-induced conformational changes. The results prove that this labeling strategy in combination with PELDOR has a great potential for studying both structure and dynamics of oligonucleotides and their complexes with various ligands.

AB - Site-directed spin labeling and pulsed electron–electron double resonance (PELDOR or DEER) have previously been applied successfully to study the structure and dynamics of nucleic acids. Spin labeling nucleic acids at specific sites requires the covalent attachment of spin labels, which involves rather complicated and laborious chemical synthesis. Here, we use a noncovalent label strategy that bypasses the covalent labeling chemistry and show that the binding specificity and efficiency are large enough to enable PELDOR or DEER measurements in DNA duplexes and a DNA duplex bound to the Lac repressor protein. In addition, the rigidity of the label not only allows resolution of the structure and dynamics of oligonucleotides but also the determination of label orientation and protein-induced conformational changes. The results prove that this labeling strategy in combination with PELDOR has a great potential for studying both structure and dynamics of oligonucleotides and their complexes with various ligands.

KW - Binding

KW - PULSED ELECTRON

KW - Nucleic-acids

KW - Conformation

KW - Lactose operon

KW - Distance measurements

KW - Operator

KW - EPR spectroscopy

KW - Electron-paramagnetic-resonance

KW - LAC repressor

U2 - 10.1093/nar/gks817

DO - 10.1093/nar/gks817

M3 - Article

SN - 0305-1048

VL - 41

JO - Nucleic Acids Research

JF - Nucleic Acids Research

IS - 1

M1 - e11

ER -

Protein-induced changes in DNA structure and dynamics observed with noncovalent site-directed spin-labeling and PELDOR

Abstract

Keywords

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Projects

Multifunctional molecular machines: multifunctional molecular machines acting on DNA

ConformationalStates of MembranceProtein: Conformational states of membrane proteins: Technology development for bioscience

ConformationalStates of MembraneProtein: Conformational states of membranes proteins: Technology development for bioscience

Cite this