Colorimetric Split G-Quadruplex Probes for Nucleic Acid Sensing: Improving Reconstituted DNAzyme's Catalytic Efficiency via Probe Remodeling
Colorimetric Split G-Quadruplex Probes for Nucleic Acid Sensing: Improving Reconstituted DNAzyme's Catalytic Efficiency via Probe Remodeling
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DOI：
10.1021/ja902951b
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发表时间：
2009-07-29
影响因子：
15
通讯作者：
Sintim, Herman O.
中科院分区：
文献类型：
--
作者：
Nakayama, Shizuka;Sintim, Herman O.
断裂的富含G的DNA探针可以在生物分析物如DNA的存在下组装成活性过氧化物酶模拟DNA酶，从而提供了检测生物样品中的分析物的简单且廉价的手段。一项旨在揭示显著的探针结构特征和反应条件的综合研究揭示了这些重要的发现：（a）在G-四链体结构中连接G3段的环可以被茎-环或环-茎-环基序取代，而不会使所得的四链体结构不稳定;赋予断裂的富含G的探针有限互补的区域导致更有效的重构酶。(b)在反平行的G-四链体DNA酶中加入氯化血红素，导致G-四链体DNA酶的CD谱发生蓝移。(c)与G-四链体结构相邻的DNA基序的结构影响重组酶的稳定性和酶促活性。(d)过量存在的单价阳离子的性质是G-四链体DNA酶的周转数的关键决定因素; G-四链体DNA酶在含有铵离子的缓冲液中的分解比含有钠或钾离子的缓冲液慢。这些发现对于设计使用过氧化物酶模拟G-四链体作为检测标记的生物测定是重要的。
Split G-rich DNA probes can assemble into active peroxidase-mimicking DNAzymes in the presence of bioanalytes such as DNA, thereby providing a simple and cheap means to detect analytes in biological samples. A comprehensive study designed to reveal the salient probe architectural features and reaction conditions that facilitate facile reconstitution into enzymatically proficient enzymes unveiled these important findings: (a) The loops that connect the G3-tracts in a G-quadruplex structure can be replaced with a stem-loop or loop-stem-loop motif without destabilizing the resulting quadruplex structure; endowing the split G-rich probes with regions of limited complementarity leads to more proficient reconstituted enzymes. (b) The addition of hemin to antiparallel G-quadruplex DNAzymes lead to a blue shift in the CD spectra of the G-quadruplex DNAzymes. (c) The architectures of the DNA motifs that lie adjacent to the G-quadruplex structure influence both the stability and the enzymatic proficiency of the reconstituted enzymes. (d) The nature of the monovalent cation that is present in excess is a key determinant of the turnovernumber of the G-quadruplex DNAzyme; decomposition of G-quadruplex DNAzymes is slower in buffers that contain ammonium ions than those that contain sodium or potassium ions. These findings are important for the design of bioassays that use peroxidase-mimicking G-quadruplexes as detection labels.