A Quantitative Raman Spectroscopic Signal for Metal-Phosphodiester Interactions in Solution
A Quantitative Raman Spectroscopic Signal for Metal-Phosphodiester Interactions in Solution
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DOI：
10.1021/bi901866u
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发表时间：
2010-04-06
期刊：
影响因子：
2.9
通讯作者：
Harris, Michael E.
中科院分区：
文献类型：
--
作者：
Christian, Eric L.;Anderson, Vernon E.;Harris, Michael E.
金属离子磷酸二酯相互作用的准确鉴定和定量对于理解金属离子作为大RNA三维折叠的决定因素以及作为RNA和蛋白质磷酸二酯酶活性位点的辅助因子的作用至关重要。由于溶液中的核酸和其他磷酸二酯形成的直接和间接相互作用的动态和复杂的混合物，实现这一目标是困难的。为了解决这个问题，拉曼光谱已被用来测量由于金属相互作用引起的键振动能的变化。然而，内层、氢键和静电相互作用对磷酰氧拉曼光谱的贡献尚未得到定量分析。在这里，我们报告所有三种形式的金属离子相互作用都会导致非桥联磷酸氧 (vsPO(2)(-)) 对称振动的拉曼信号衰减，而只有内球配位会导致溶液中 vsPO(2)(-) 明显转变为更高波数 (v(s)PO(2)(-) M）。 vsPO(2)(-)M 的形成取决于金属离子特性和位点特异性金属离子结合的准确测量。此外，反映vsPO(2)(-)和vsPO(2)(-)M (Delta vM)之间能量差异的光谱参数在很大程度上对磷酸二酯结构的变化不敏感，但强烈依赖于相互作用的金属离子的绝对电负性和硬度。总之，这些研究为使用 vsPO(2)(-)M 和 Delta vM 作为通用光谱特征来定量分析金属结合亲和力和鉴定与溶液中磷酸二酯相关的金属离子提供了强有力的实验支持。
Accurate identification and quantification of metal ion phosphodiester interactions are essential for understanding the role of metal ions as determinants of three-dimensional folding of large RNAs and as cofactors in the active sites of both RNA and protein phosphodiesterases. Accomplishing this goal is difficult due to the dynamic and complex mixture of direct and indirect interactions formed with nucleic acids and other phosphodiesters in solution. To address this issue, Raman spectroscopy has been used to measure changes in bond vibrational energies due to metal interactions. However, the contributions of inner-sphere, H-bonding, and electrostatic interactions to the Raman spectrum of phosphoryl oxygens have not been analyzed quantitatively. Here, we report that all three forms of metal ion interaction result in attenuation of the Raman signal for the symmetric vibration of the nonbridging phosphate oxygens (vsPO(2)(-)), while only inner-sphere coordination gives rise to an apparent shift of vsPO(2)(-) to higher wavenumbers (v(s)PO(2)(-) M) in solution. Formation of vsPO(2)(-)M is shown to be both dependent on metal ion identity and an accurate measure of site-specific metal ion binding. In addition, the spectroscopic parameter reflecting the energetic difference between vsPO(2)(-) and vsPO(2)(-)M (Delta vM) is largely insensitive to changes in phosphodiester structure but strongly dependent on the absolute electronegativity and hardness of the interacting metal ion. Together, these studies provide strong experimental support for the use of vsPO(2)(-)M and Delta vM as general spectroscopic features for the quantitative analysis of metal binding affinity and the identification of metal ions associated with phosphodiesters in solution.