Smirnov I, Shafer R H
Department of Pharmaceutical Chemistry, School of Pharmacy, University of California at San Francisco, San Francisco, CA 94143-0446, USA.
J Mol Biol. 2000 Feb 11;296(1):1-5. doi: 10.1006/jmbi.1999.3441.
DNA quadruplex structures based on the guanine quartet are typically stabilized by monovalent cations such as K(+), Na(+), or NH(+)(3). Certain divalent cations can also induce quadruplex formation, such as Sr(2+). Here we show that Pb(2+) binds with unusually high affinity to the thrombin binding aptamer, d(GGTTGGTGTGGTTGG), inducing a unimolecular folded structure. At micromolar concentrations the binding is stoichiometric, and a single lead cation suffices to fold the aptamer. The lead-induced changes in UV and CD spectra are characteristic of folded quadruplexes, although the long wavelength CD maximum occurs at 312 nm rather than the typical value of 293 nm. The one-dimensional exchangeable proton NMR spectrum shows resonances expected for imino protons involved in guanine quartet base-pairing. Furthermore, two-dimensional NMR experiments reveal NOE contacts typically seen in folded structures formed by guanine quartets, such as the K(+) form of the thrombin aptamer. Only sequences capable of forming guanine quartets appear to bind Pb(+2) tightly and change conformation. This sequence-specific, tight DNA binding may be relevant to possible genotoxic effects of lead in the environment.
基于鸟嘌呤四联体的DNA四重结构通常由单价阳离子（如K⁺、Na⁺或NH₄⁺）稳定。某些二价阳离子也能诱导四重结构形成，比如Sr²⁺。在此我们表明，Pb²⁺以异常高的亲和力与凝血酶结合适体d(GGTTGGTGTGGTTGG)结合，诱导形成单分子折叠结构。在微摩尔浓度下，结合是化学计量的，单个铅离子就足以使适体折叠。铅诱导的紫外和圆二色光谱变化是折叠四重结构的特征，尽管长波长圆二色光谱最大值出现在312 nm而非典型的293 nm。一维可交换质子核磁共振谱显示出参与鸟嘌呤四联体碱基配对的亚氨基质子预期的共振峰。此外，二维核磁共振实验揭示了通常在由鸟嘌呤四联体形成的折叠结构（如凝血酶适体的K⁺形式）中出现的核Overhauser效应（NOE）接触。只有能够形成鸟嘌呤四联体的序列似乎能紧密结合Pb²⁺并改变构象。这种序列特异性的紧密DNA结合可能与环境中铅可能的遗传毒性效应有关。
