SUNY at Albany
June 19-23, 2001
Internal Dynamics in a Site-Specific 15N-Labeled DNA Triple Helix
Triple-helical DNA structures are of current interest as potential ways to block protein binding to DNA and to chemically modify DNA in a sequence specific manner. Formation of triple helical structures involves Hoogsteen hydrogen bonds between bases in the third strand and selected sites in the major groove of the DNA double helix. The exocyclic amino group of adenine is one of the key sites for this Hoogsteen hydrogen bonding in TAT, AAT, and A+GC triads.
In the present work, we have used 1H-15N heteronuclear NMR spectroscopy to characterize the stability and the dynamics of the amino group of adenine in the DNA triple helix formed by the 31-mer oligonucleotide 5'-d(AGAGAGAA-CCCC-TTCTCTCT-TTT-TCTCTCTT)-3'. Previous NMR studies by Feigon and coworkers (1) have shown that this triple helix contains canonical TAT and C+GC base triads. The energetics of each of these triads has been previously characterized by this laboratory (2). The adenine of interest to the present work (highlighted in the sequence) was labeled with 15N at its N6 position, and the NMR resonances of the two amino protons were selectively observed by 15N-HSQC-editing. The rates of rotation of the adenine amino group were measured in HSQC-edited transfer of magnetization experiments. The activation parameters for the rotational motion were determined from the temperature dependence of the rotation rates in the range from 1 to 45¼C. The energetics of the rotation of the adenine amino group in this DNA triple helix will be compared to that previously determined by this laboratory for a DNA double-helix (3).
Supported by a grant from the National Science FoundationReferences and Footnotes
Lihong Jiang (1), and Irina M. Russu (2),
Department of Molecular Biology and Biochemistry (1), and Department of Chemistry (2), Molecular Biophysics Program, Wesleyan University, Middletown, CT 06459, USA.