Ramreddy T, Rao B J, Krishnamoorthy G
Department of Chemical Science and Department of Biological Sciences, Tata Institute of Fundamental Research, Homi Bhabha Road, Mumbai 400005, India.
J Phys Chem B. 2007 May 24;111(20):5757-66. doi: 10.1021/jp068818f. Epub 2007 May 1.
It is well recognized that structure and dynamics of DNA strands guide proteins toward their cognate sites in DNA. While the dynamics is controlled primarily by the nucleotide sequence, the context of a particular sequence in relation to an open end could also play a significant role. In this work we have used the fluorescent analogue of adenine, 2-aminopurine (2-AP), to extract information on site-specific dynamics of DNA strands associated with 30-70 nucleotides length. Measurement of fluorescence lifetime and anisotropy decay kinetics in various types of DNA strands in which 2-AP was located in specific positions revealed novel insights into the dynamics of strands. We find that in single-stranded (ss) DNA, the extent of motional dynamics of the bases falls off sharply from the very end toward the middle of the strand. In contrast, the flexibility of the backbone decreases more gradually in the same direction. In double-stranded (ds) DNA, the level of base-pair fraying increases toward the ends in a graded manner. Surprisingly, the same is countered by the presence of ss-overhangs emanating from dsDNA ends. Moreover, the extent of concerted motion of bases in duplex DNA increased from the end to the middle of the duplex, a result which is both striking and counterintuitive. Most surprisingly, the two complementary strands of a duplex that were unequal in length exhibited differential dynamics: the longer one with overhangs showed a distinctly higher level of flexibility than the recessed shorter strand in the same duplex. All these results, taken together, provoke newer insights in our understanding of how different bases in DNA strands are endowed with specific dynamic properties as a function of their positions. These properties are likely to be used in facilitating specific recognitions of DNA bases by proteins during various DNA-protein interaction systems.
人们普遍认识到，DNA链的结构和动力学引导蛋白质趋向其在DNA中的同源位点。虽然动力学主要由核苷酸序列控制，但特定序列相对于开放末端的背景也可能发挥重要作用。在这项工作中，我们使用了腺嘌呤的荧光类似物2-氨基嘌呤（2-AP）来获取与30-70个核苷酸长度相关的DNA链位点特异性动力学信息。测量2-AP位于特定位置的各种类型DNA链中的荧光寿命和各向异性衰减动力学，揭示了链动力学的新见解。我们发现，在单链（ss）DNA中，碱基的运动动力学程度从链的末端向中间急剧下降。相比之下，主链的柔韧性在同一方向上下降得更为缓慢。在双链（ds）DNA中，碱基对解链的水平朝着末端呈梯度增加。令人惊讶的是，dsDNA末端产生的ss悬垂会抵消这种情况。此外，双链DNA中碱基协同运动的程度从双链的末端向中间增加，这一结果既引人注目又违反直觉。最令人惊讶的是，长度不等的双链的两条互补链表现出不同的动力学：带有悬垂的较长链显示出比同一双链中凹陷的较短链明显更高的柔韧性水平。所有这些结果综合起来，引发了我们对DNA链中不同碱基如何根据其位置赋予特定动态特性的新认识。这些特性可能用于在各种DNA-蛋白质相互作用系统中促进蛋白质对DNA碱基的特异性识别。
