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Errors in RNA NOESY Distance Measurements in Chimeric and Hybrid Duplexes: Differences in RNA and DNA Proton Relaxation

机译：嵌合和杂交双链体中RNA NOESY距离测量中的错误：RNA和DNA质子弛豫的差异

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Nuclear magnetic resonance experiments reveal that the base H8/H6 protons of oligoribo-nucleotides (RNA) have T1 relaxation times that are distinctly longer than those of oligodeoxyribonucleotides (DNA). Similarly, the T1 values for the RNA H1′ protons are approximately twice those of the corresponding DNA H1′ protons. These relaxation differences persist in single duplexes containing covalently linked RNA and DNA segments and cause serious overestimation of distances involving RNA protons in typical NOESY spectra collected with a duty cycle of 2–3 s. NMR and circular dichroism experiments indicate that the segments of RNA maintain their A-form geometry even in the interior of DNA–RNA–DNA chimeric duplexes, suggesting that the relaxation times are correlated with the type of helix topology. The difference in local proton density is the major cause of the longer nonselective T1s of RNA compared to DNA, although small differences in internal motion cannot be completely ruled out. Fortunately, any internal motion differences that might exist are shown to be too small to affect cross-relaxation rates, and therefore reliable distance data can be obtained from time-dependent NOESY data sets provided an adequately long relaxation delay is used. In hybrid or chimeric RNA–DNA duplexes, if the longer RNA relaxation times are not taken into account in the recycle delay of NOESY pulse sequences, serious errors in measuring RNA proton distances are introduced.
机译：核磁共振实验表明，寡核糖核苷酸（RNA）的碱基H8 / H6质子具有的T1弛豫时间明显长于寡脱氧核糖核苷酸（DNA）的T1弛豫时间。同样，RNA H1'质子的T1值大约是相应DNA H1'质子的T1值的两倍。这些松弛差异在包含共价连接的RNA和DNA片段的双链体中仍然存在，并导致在2到3 s的占空比收集的典型NOESY光谱中，涉及RNA质子的距离被严重高估。 NMR和圆二色性实验表明，即使在DNA–RNA–DNA嵌合双链体内部，RNA片段仍保持其A型几何结构，这表明弛豫时间与螺旋拓扑的类型相关。尽管不能完全排除内部运动的微小差异，但局部质子密度的差异是RNA与DNA相比更长的非选择性T1的主要原因。幸运的是，任何可能存在的内部运动差异都显示为太小而无法影响交叉松弛率，因此，只要使用足够长的弛豫延迟，就可以从时间相关的NOESY数据集中获得可靠的距离数据。在杂合或嵌合的RNA-DNA双链体中，如果在NOESY脉冲序列的循环延迟中未考虑较长的RNA弛豫时间，则会在测量RNA质子距离时引入严重的误差。

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