Preparation, resonance assignment, and preliminary dynamics characterization of residue specific 13C/15N-labeled elongated DNA for the study of sequence-directed dynamics by NMR

Nikolova, Evgenia N.; Al‐Hashimi, Hashim M.

doi:10.1007/s10858-009-9350-y

Cited by 16 publications

(17 citation statements)

References 71 publications

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“…This suggests the presence of sugar-backbone fluctuations occurring at ns timescales that are masked by overall rotational diffusion; lowering the temperature decouples the two motions by slowing down overall diffusion, allowing better resolution of the local dynamics. 66,67 …”

Section: Resultsmentioning

confidence: 99%

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Probing Sequence-Specific DNA Flexibility in A-Tracts and Pyrimidine-Purine Steps by Nuclear Magnetic Resonance ¹³C Relaxation and Molecular Dynamics Simulations

et al. 2012

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Sequence-specific DNA flexibility plays a key role in a variety of cellular interactions that are critical for gene packaging, expression, and regulation. Yet, few studies have experimentally explored the sequence dependence of DNA dynamics that occur on biologically relevant timescales. Here, we use nuclear magnetic resonance (NMR) carbon spin relaxation combined with molecular dynamics (MD) simulations to examine the picosecond to nanosecond dynamics in a variety of dinucleotide steps as well as in varying length homopolymeric An•Tn repeats (An-tracts, n = 2, 4 and 6) that exhibit unusual structural and mechanical properties. We extend the NMR spin relaxation timescale sensitivity deeper into the nanosecond regime by using glycerol and a longer DNA duplex to slow down overall tumbling. Our studies reveal a structurally unique A-tract core (for n > 3) that is uniformly rigid, flanked by junction steps that show increasing sugar flexibility with A-tract length. High sugar mobility is observed at pyrimidine residues at the A-tract junctions, which is encoded at the dinucleotide level (CA, TG and CG steps) and increases with A-tract length. The MD simulations reproduce many of these trends, particularly the overall rigidity of A-tract base and sugar sites, and suggest that the sugar-backbone dynamics could involve transitions in sugar pucker and phosphate backbone BI↔BII equilibria. Our results reinforce an emerging view that sequence-specific DNA flexibility can be imprinted in dynamics occurring deep within the nanosecond time regime that is difficult to characterize experimentally at the atomic level. Such large amplitude sequence-dependent backbone fluctuations might flag the genome for specific DNA recognition.

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Section: Resultsmentioning

confidence: 99%

“…We cannot rule out large-scale helical bending motions as the source for the enhanced backbone dynamics at A-tract junctions, even though such motions could not be previously detected using a DNA domain-elongation approach. 67 …”

Section: Discussionmentioning

confidence: 99%

Probing Sequence-Specific DNA Flexibility in A-Tracts and Pyrimidine-Purine Steps by Nuclear Magnetic Resonance ¹³C Relaxation and Molecular Dynamics Simulations

et al. 2012

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“…The authors noted the potential difficulties posed by 13 C– 13 C scalar and dipolar interactions for C6 of pyrimidines and other 13 C sites in fully labeled RNA molecules. As for proteins, isotopic labeling strategies have been developed to increase the number of isolated 13 C spins suitable for relaxation dispersion and other relaxation measurements [110-112]. Selective excitation and decoupling schemes can be used to minimize complications of 13 C– 13 C interactions in natural abundance and isotopically enriched molecules [113].…”

Section: Applications To Nucleic Acidsmentioning

confidence: 99%

Chemical exchange in biomacromolecules: Past, present, and future

Palmer

2014

Journal of Magnetic Resonance

239

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The perspective reviews quantitative investigations of chemical exchange phenomena in proteins and other biological macromolecules using NMR spectroscopy, particularly relaxation dispersion methods. The emphasis is on techniques and applications that quantify the populations, interconversion kinetics, and structural features of sparsely populated conformational states in equilibrium with a highly populated ground state. Applications to folding, mol ecular recognition, catalysis, and allostery by proteins and nucleic acids are highlighted.

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“…Importantly, we often observed extensive mobility at highly conserved residues that are known to undergo functionally conformational rearrangements during recognition and catalysis. In later studies, we also adapted the domain-elongation strategy to study DNA dynamics and this led us to uncover nanosecond end-fraying motions that penetrate deep into the helix interior [54]. …”

Section: Picosecond-to-nanosecond Motions In Rna Using Domain-elongatmentioning

confidence: 99%

NMR studies of nucleic acid dynamics

Al‐Hashimi

2013

Journal of Magnetic Resonance

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Nucleic acid structures have to satisfy two diametrically opposite requirements; on one hand they have to adopt well-defined 3D structures that can be specifically recognized by proteins; on the other hand, their structures must be sufficiently flexible to undergo very large conformational changes that are required during key biochemical processes, including replication, transcription, and translation. How do nucleic acids introduce flexibility into their 3D structure without losing biological specificity? Here, I describe the development and application of NMR spectroscopic techniques in my laboratory for characterizing the dynamic properties of nucleic acids that tightly integrate a broad set of NMR measurements, including residual dipolar couplings, spin relaxation, and relaxation dispersion with sample engineering and computational approaches. This approach allowed us to obtain fundamental new insights into directional flexibility in nucleic acids that enable their structures to change in a very specific functional manner.

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Preparation, resonance assignment, and preliminary dynamics characterization of residue specific 13C/15N-labeled elongated DNA for the study of sequence-directed dynamics by NMR

Cited by 16 publications

References 71 publications

Probing Sequence-Specific DNA Flexibility in A-Tracts and Pyrimidine-Purine Steps by Nuclear Magnetic Resonance ¹³C Relaxation and Molecular Dynamics Simulations

Probing Sequence-Specific DNA Flexibility in A-Tracts and Pyrimidine-Purine Steps by Nuclear Magnetic Resonance ¹³C Relaxation and Molecular Dynamics Simulations

Chemical exchange in biomacromolecules: Past, present, and future

NMR studies of nucleic acid dynamics

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Preparation, resonance assignment, and preliminary dynamics characterization of residue specific 13C/15N-labeled elongated DNA for the study of sequence-directed dynamics by NMR

Cited by 16 publications

References 71 publications

Probing Sequence-Specific DNA Flexibility in A-Tracts and Pyrimidine-Purine Steps by Nuclear Magnetic Resonance 13C Relaxation and Molecular Dynamics Simulations

Probing Sequence-Specific DNA Flexibility in A-Tracts and Pyrimidine-Purine Steps by Nuclear Magnetic Resonance 13C Relaxation and Molecular Dynamics Simulations

Chemical exchange in biomacromolecules: Past, present, and future

NMR studies of nucleic acid dynamics

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Probing Sequence-Specific DNA Flexibility in A-Tracts and Pyrimidine-Purine Steps by Nuclear Magnetic Resonance ¹³C Relaxation and Molecular Dynamics Simulations

Probing Sequence-Specific DNA Flexibility in A-Tracts and Pyrimidine-Purine Steps by Nuclear Magnetic Resonance ¹³C Relaxation and Molecular Dynamics Simulations