Effect of base-pair sequence on the conformations and thermally induced transitions in oligodeoxyribonucleotides containing only AT base pairs

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Abstract
Tm curves, CD spectra, and kinetics results of the self-complementary DNA dodecamers d(A6T6), d(A3T3A3T3), d(A2T2A2T2A2T2), d(ATATATATATAT), and d(T6A6) demonstrate that the thermal transitions of these oligomers at low salt concentration involve a hairpin intermediate. At high salt concentrations (> 0.1 M Na+) only a duplex to denatured-strand transition appears to occur. The temperature and salt-concentration regions of the transitions are very sequence dependent. Alternating-type AT sequences have a lower duplex stability and a greater tendency to form hairpins than sequences containing more nonalternating AT base pairs. Of the two nonalternating sequences, d(T6A6) is significantly less table than d(A6T6). Both oligomers have CD curves that are very similar to the unusual CD spectrum of poly(dA).cntdot.poly(dT). The Raman spectra of these two oligomers are also quite similar, but at low temperature, small intensity differences in two backbone modes and three nucleotide vibrations are obtained. The hairpin to duplex transition for the AT dodecamers was examined by salt-jump kinetics measurements. The transition is faster than transitions for palindromic-sequence containing terminal GC base pairs. Stopped-flow kinetics studies indicate that the transition is second order and has a relatively low activation energy. The reaction rate increases with increasing ionic strength. These results are consistent with a three-step mechanism for the hairpin to duplex reaction: (i) fraying of the hairpin oligomers'' terminal base pairs, (ii) a rate-determining bimolecular step involving formation of a cruciform-type intermediate from two hairpin oligomers will open terminal base pairs, and (iii) base-pairs migration and formation in the intermediate to give the duplex.