Preparation, resonance assignment, and preliminary dynamics characterization of residue specific 13C/15N-labeled elongated DNA for the study of sequence-directed dynamics by NMR.
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 (13)C/(15)N-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.
Nikolova
Evgenia N
EN
Department of Chemistry, Biophysics and Chemical Biology Program, University of Michigan, 930 North University Avenue, Ann Arbor, MI 48109-1055, USA.
Al-Hashimi
Hashim M
HM
D001483:Base Sequence; D002247:Carbon Isotopes / Q000737:chemistry; D004247:DNA / Q000737:chemistry*; D008954:Models, Biological; D008969:Molecular Sequence Data; D009587:Nitrogen Isotopes / Q000737:chemistry; D019906:Nuclear Magnetic Resonance, Biomolecular / Q000379:methods*; D016133:Polymerase Chain Reaction; D017422:Sequence Analysis, DNA / Q000379:methods*
D002247:Carbon Isotopes; D009587:Nitrogen Isotopes; D004247:DNA