Prototropically Allosteric Probe for Superbly Selective DNA Analysis
Prototropically Allosteric Probe for Superbly Selective DNA Analysis
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用于高选择性 DNA 分析的质子变构探针
DOI：
10.1021/acs.analchem.7b02077
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发表时间：
2017
影响因子：
7.4
通讯作者：
Xu Zhiai
中科院分区：
文献类型：
--
作者：
Lin Fan;Zhou Yufeng;Li Qiusha;Zhou Xiaoshun;Shao Yong;Habermeyer Benoit;Wang Hui;Shi Xinghua;Xu Zhiai
生物传感器进化的选择性核苷酸识别需要针对结合模式敏感的读出进行合理的探针设计，但不会严重损害上下文序列的选择性。在这项工作中，我们合成了一种双功能（三羟苯基）卟啉（POH3），以三羟苯基取代基和四吡咯大环分别作为识别单元（RU）和荧光信号单元（SU）来靶向双链DNA中的脱碱基位点（AP位点）。 RU 和 SU 彼此分离，但具有原变性变构。我们发现适当的 pH 值有利于 POH3 的非荧光奎宁/吡咯 (O-NH) 构象异构体的形成。然而，O-NH 中 RU 与 AP 位点对面的靶胞嘧啶的互补氢键通过向苯酚/异吡咯 (OH-N) 构象异构体的质子变构打开 SU 荧光，而碱基胸腺嘧啶、鸟嘌呤和腺嘌呤完全沉默这种变构，表明单核苷酸多态性 (SNP) 分析具有极好的选择性。使用具有其他羟基取代基模式的卟啉也证明了原变变构在实现这种 SNP 选择性中的作用。由于SU与RU分离，SU不直接参与与AP位点的相互作用，因此，对于侧翼具有鸟嘌呤（DNA中最容易氧化的碱基）的DNA也实现了开启选择性。这种对侧翼碱基身份的耐受性在以前的研究中很少实现。此外，其他DNA结构不能带来这种变构，这表明AP位点设计和质子变构探针的组合配方将在基于DNA的传感器中得到广泛的应用。
Selective nucleotide recognition for biosensor evolution requires rational probe design toward the binding-pattern-susceptible readout but without serious poison in selectivity from the context sequences. In this work, we synthesized a dual-function (trihydroxyphenyl)porphyrin (POH3) to target the abasic site (AP site) in ds-DNA using the trihydroxyphenyl substituent and the tetrapyrrole macrocycle as the recognition unit (RU) and the fluorescent signal unit (SU), respectively. RU and SU are separated from each other but are prototropically allosteric. We found that an appropriate pH favors formation of the nonfluorescent quinine/pyrrole (O–NH) conformer of POH3. However, the complementary hydrogen bonding of RU in O–NH with the target cytosine opposite the AP site switches on the SU fluorescence through prototropic allostery toward the phenol/isopyrrole (OH–N) conformer, while the bases thymine, guanine, and adenine totally silence this allostery, suggesting a superb selectivity in single-nucleotide polymorphism (SNP) analysis. The role of the prototropic allostery in achieving such SNP selectivity is also evidenced using porphyrins with other hydroxyl substituent patterns. Because of the SU separation from RU, SU is not directly involved in the interaction with the AP site, and thus, the turn-on selectivity is also realized for DNA with flanking guanine, the most easily oxidized base in DNA. This tolerance to the flanking base identity has seldom been achieved in previous studies. Additionally, other DNA structures cannot bring this allostery, indicating that the combination recipe of the AP site design and the prototropically allosteric probe will find wide applications in DNA-based sensors.