| Abstrakt: |
Lefevre et al. originally observed conformational transitions at the TpA step in the TTAA Pribnow box sequence of the trp promoter [Lefevre, J.-F., Lane, A. N., & Jardetzky, O. (1985) FEBS Lett. 190, 37-40]. In 500-MHz 1H NMR studies on the TnA(n)-containing DNA oligonucleotides [d(CGAGGTTTAAACCTCG)]2, [d(GCTCCTTTAAAGGAGC)]2, and [d(GCCGTTAACGGC)]2, we observe that, in addition to the H2 proton (which resides in the minor groove of DNA), the H8 proton of the first adenine (which resides in the major groove) is also broadened due to motion at the TpA junction. In analogous 16-mers where the T3A3 segment has been replaced by an A3T3 sequence, and therefore contains CA, GA, and AT steps (but no TA steps), all adenine proton resonances are narrow, indicating that the broadening occurs only at TpA steps. Assuming chemical exchange in the form of conformational dynamics, e.g., oscillation of the purine base about the glycosidic torsion angle, the experimental 500-MHz 1H T1 rho and 2D-NOESY data were used to constrain the correlation time of the internal motion to a range between the T1 and T1 rho minima. Calculated line shapes using a two-site exchange model indicate that the motion has an amplitude of 20-50 degrees with an associated tau c of ca. 1.6 x 10(-4) to 1.0 x 10(-5) s rad-1, respectively. The mobility appears to be a consequence of the structure at the TpA junction which is characterized by (1) a wide minor groove between two regions of narrow minor groove, (2) an unusual average orientation of the adenine heterocycle probably resulting from a poor base-stacking interaction of the adenine with the preceding thymine, and (3) a sharp discontinuity in the sugar conformation at the TpA step. |
