Solution structure of an A-tract DNA bend

The solution structure of a DNA dodecamer d(GGCAAAAAACGG)/d(CCGTTTTTTGCC) c ontaining an A-tract has been determined by NMR spectroscopy with residual dipolar couplings. The structure shows an overall helix axis bend of 19 deg rees in a geometry consistent with solution and gel electrophoresis experim ents. 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-pai rs. 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 tw ist 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 geometr y of the A-tract different from all X-ray structures, the proton on N6 of a denine 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 jun ction between the GC flank and the A-tract through a combination of tilt an d 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 o f 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 degr ees inclination. This bend is a combination of tilt and roll. The large cha nge in inclination allows the formation of a hydrogen bond between the prot ons of N4 of the 3' cytosine and the O6 of the next 3' base, a guanine, sta bilizing the roll component in the bend. These structural features differ f rom 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 o f the A-tract. This structure has no negative inclination in most of the ba ses 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 electroph oresis experiments. (C) 2001 Academic Press.