Abstract
DNA is a highly flexible molecule that undergoes functionally important structural transitions in response to external cellular stimuli. Atomic level spin relaxation NMR studies of DNA dynamics have been limited to short duplexes in which sensitivity to biologically relevant fluctuations occurring at nanosecond timescales is often inadequate. Here, we introduce a method for preparing residue-specific 13C/15N-labeled elongated DNA along with a strategy for establishing resonance assignments and apply the approach to probe fast inter-helical bending motions induced by an adenine tract. Preliminary results suggest the presence of elevated A-tract independent end-fraying internal motions occurring at nanosecond timescales, which evade detection in short DNA constructs and that penetrate deep (7 bp) within the DNA helix and gradually fade away towards the helix interior.
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Acknowledgments
We thank Dr. Alexandar L. Hansen for stimulating discussions and for critically reading the manuscript and Dr. Alexander V. Kurochkin for maintenance of the NMR instruments. We gratefully acknowledge the Michigan Economic Development Cooperation and the Michigan Technology Tri-Corridor for support in the purchase of a 600 MHz spectrometer. This work was supported by a National Science Foundation CAREER award (MCB 0644278) received by H.M.A. E.N.N. was supported by a Rackham International Fellowship awarded by the University of Michigan.
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Nikolova, E.N., Al-Hashimi, H.M. Preparation, resonance assignment, and preliminary dynamics characterization of residue specific 13C/15N-labeled elongated DNA for the study of sequence-directed dynamics by NMR. J Biomol NMR 45, 9–16 (2009). https://doi.org/10.1007/s10858-009-9350-y
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DOI: https://doi.org/10.1007/s10858-009-9350-y