Solution structure of an A-tract DNA bend

The solution structure of a DNA dodecamer d(GGCAAAAAACGG)/d(CCGTTTTTTGCC) containing an A-tract has been determined by NMR spectroscopy with residual dipolar couplings. The structure shows an overall helix axis bend of 19 degrees in a geometry consistent with solution and gel electrophoresis experiments. Fourteen degrees of the bending occurs in the GC regions flanking the A-tract. The remaining 5 degrees is spread evenly over its six AT base-pairs. The A-tract is characterized by decreasing minor groove width from the 5' to the 3' direction along the A strand. This is a result of propeller twist in the AT pairs and the increasing negative inclination of the adenine bases at the 3' side of the run of adenine bases. The four central thymine bases all have negative inclination throughout the A-tract with an average value of -6.1 degrees. Although this negative inclination makes the geometry of the A-tract different from all X-ray structures, the proton on N6 of adenine and the O4 of thymine one step down the helix are within distance to form bifurcated hydrogen bonds. The 5' bend of 4 degrees occurs at the junction between the GC flank and the A-tract through a combination of tilt and roll. The larger 3' bend, 10 degrees, occurs in two base steps: the first composed of tilt, -4.1 degrees, and the second a combination of tilt, -4.2 degrees, and roll, 6.0 degrees. This second step is a direct consequence of the change in inclination between an adjacent cytosine base, which has an inclination of -12 degrees, and the next base, a guanine, which has 3 degrees inclination. This bend is a combination of tilt and roll. The large change in inclination allows the formation of a hydrogen bond between the protons of N4 of the 3' cytosine and the O6 of the next 3' base, a guanine, stabilizing the roll component in the bend. These structural features differ from existing models for A-tract bends. For comparison, we also determined the structure of the control sequence, d(GGCAAGAAACGC)/d(CCGTTTCTTGCC), with an AT to GC transition in the center of the A-tract. This structure has no negative inclination in most of the bases within the A-tract, resulting in a bend of only 9 degrees. When ligated in phase, the control sequence has nearly normal mobility in gel electrophoresis experiments. (C) 2001 Academic Press.