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通过荧光和荧光共振能量转移研究研究 14mer DNA 寡核苷酸与 CTAB 之间的相互作用。

An investigation on interaction between 14mer DNA oligonucleotide and CTAB by fluorescence and fluorescence resonance energy transfer studies.

机构信息

Institute for Genomics and Integrative Biology, CSIR, Mall Road, Delhi 110 007, India.

出版信息

J Phys Chem B. 2010 Jun 10;114(22):7602-8. doi: 10.1021/jp909522r.

Abstract

Possible interaction mechanisms between oligonucleotide (DNA) of 14 base pairs with cetyl trimethyl ammonium bromide (CTAB) were postulated based on fluorescence and fluorescence resonance energy transfer (FRET) studies. Detailed FRET investigations were carried out by fluorometric titrations of the surfactant with various oligonucleotide duplexes with 5'-tagged fluorescein (donor) (D(D)), 5'-tagged TAMRA (acceptor) (D(A)) and both (D(DA)). In general, fluorescence spectra of the duplexes (D(D), D(A) and D(DA)) revealed a reduction in the fluorescence intensities of 5'-fluorescein as well as 5'-TAMRA and thereafter an attainment of saturation with increase in the surfactant concentration. The observed changes in the oligonucleotide fluorescence intensities for the duplexes under investigation could be attributed to the microenvironmental changes during the oligonucleotide-CTAB interaction. Considering together, it appeared that the interaction is a three-stage process, wherein the initial addition of surfactant caused neutralization of the 14mer at Z(+/-)(1) = 0.8, which is manifested by a slight reduction in fluorescence intensity. Further, addition of the surfactant molecules sharply reduced the fluorescence intensity of the oligonucleotide depicting oligonucleotide induced self-assembly until the second break point (Z(+/-)(2) = 1.7). From the second break point, a striking resonance energy transfer was observed from donor to acceptor, which revealed shortening of distance between 5' ends of the oligonucleotides that attained a saturation at Z(+/-)(3) = 2.5. Similar three-stage interaction of oligonucleotide with the surfactant has also been observed through fluorometric titrations in the presence of NaCl. However, in the presence of the salt, neutralization of oligonucleotide, surfactant aggregation and FRET occurred at higher charge ratios due to the screening effect of Na+ ions followed by an increase in critical association concentration (CAC) of the surfactant. Overall, investigations probe possible structural changes in the 14mer oligonucleotide-CTAB complex upon increase in the surfactant concentration.

摘要

基于荧光和荧光共振能量转移(FRET)研究,提出了 14 个碱基的寡核苷酸(DNA)与十六烷基三甲基溴化铵(CTAB)之间可能的相互作用机制。通过荧光滴定法,详细研究了表面活性剂与带有 5'标记荧光素(供体)(D(D))、5'标记 TAMRA(受体)(D(A))和两者(D(DA))的各种寡核苷酸双链体的相互作用。一般来说,双链体(D(D)、D(A)和 D(DA))的荧光光谱显示 5'-荧光素和 5'-TAMRA 的荧光强度降低,随后随着表面活性剂浓度的增加达到饱和。对于所研究的双链体,寡核苷酸荧光强度的观察到的变化可以归因于寡核苷酸-CTAB 相互作用期间微环境的变化。总的来说,似乎相互作用是一个三阶段的过程,其中表面活性剂的初始添加导致 14mer 在 Z(+/-)(1) = 0.8 处中和,这表现为荧光强度略有降低。此外,表面活性剂分子的进一步添加急剧降低了寡核苷酸的荧光强度,表明寡核苷酸诱导自组装,直到第二个断点(Z(+/-)(2) = 1.7)。从第二个断点开始,观察到从供体到受体的显著共振能量转移,这表明寡核苷酸的 5'末端之间的距离缩短,在 Z(+/-)(3) = 2.5 时达到饱和。在存在 NaCl 的情况下,通过荧光滴定也观察到寡核苷酸与表面活性剂的类似三阶段相互作用。然而,在存在盐的情况下,由于 Na+离子的屏蔽作用,寡核苷酸的中和、表面活性剂的聚集和 FRET 发生在更高的电荷比,随后表面活性剂的临界缔合浓度(CAC)增加。总的来说,研究探测了表面活性剂浓度增加时 14 mer 寡核苷酸-CTAB 复合物中可能的结构变化。

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