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The dynamics of self-complementary DNA decamers containing chemically-modified recognition sequences of the Eco RI endonuclease have been investigated by temperature-dependent picosecond time-resolved fluorescence spectroscopy. The unmodified decamer, d[CTGAATTCAG], as well as the decamer with 2-aminopurine (2AP) replacing adenine in position 5, have been shown to be B-type duplexes by 2D NMR and molecular dynamics (MD) simulations. Fluorescence anisotropy decay and MD of the 2AP-containing decamer show rapid motion of the base on timescales of 10-11 to 10-10 s. The multi-exponential fluorescence decay of the fluorescence and its temperature dependence, together with the 10-ns singleexponential decay of the isolated 2AP base, suggest that the 2AP base exists in 4 or more conformational states. All of these states appear to interconvert, but only two on the timescale of the fluorescence decay. The decamer with 1-((beta)-D-2'-deoxyribosyl)-2-pyrimidinone (dK) replacing dC in position 8 is a duplex which melts at about 21° C and shows multi-exponential fluorescence decay. Fluorescence is dominated, however, by a temperature-dependent 150-200 ps decay component accounting for >90% of the decay process. The unnormalized amplitude of this component decreases with decreasing temperature, reflecting hypochromism of dK as it stacks with its neighbors. In contrast with the other two modified bases, the isolated dK base has an extremely short fluorescence lifetime in aqueous solution, about 250 ps. The decamer with the 5-methyl derivative of dK (dS) placed in position 7 appears to be single-stranded above 10° C. Fluorescence from this decamer is thermodynamically and kinetically simpler than that from the duplexes. The decay time of the isolated d5 base is about 4.0 ns, while the decamer shows a temperature-independent 4.0 +/- 0.1 ns and shorter-lived, temperature-dependent decay components. Analysis of the data shows that completely unstacked (solvent-exposed) and partially stacked states exist. The completely unstacked state is a small component at all observed temperatures.
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