分子内裂分G-四链体脱氧核糖核酸酶用于Hg2+的特异性定量检测

G-四链体DNA酶是由核酸G-四链体与氯化血红素(Hemin)结合后形成的一种具有过氧化物酶活性的人工酶,利用这种DNA酶,可进行多种化学及生物传感器的设计。为提高G-四链体DNA酶类Hg2+传感器的选择性,本研究在传感器的设计过程中引入了分子内裂分G-四链体,即将形成G-四链体的富G序列拆分成两部分,分别放置在Hg2+探测序列的两端。在无Hg2+存在时,部分富G序列被包埋在某一分子内二倍体结构中,无法形成G-四链体。而在Hg2+存在下,Hg2+对T-T碱基错配的稳定能力可以促使Hg2+探测序列形成分子内二倍体结构,并伴随着原有分子间二倍体结构的破坏及分子内裂分G-四链体的生成。利用生成的裂分G-四链体与Hemin作用后检测体系酶活性的提高,实现Hg2+传感器的设计。利用该传感器,可在50～500 nmol/L及2.0～7.5μmol/L两个浓度范围内实现Hg2+的定量检测,检出限为47 nmol/L。由于裂分G-四链体DNA酶的使用强化了传感器对Hg2+的依赖性,极大地提高了设计的Hg2+传感器的选择性。对实际水样的加标回收结果显示,回收率为97.5%～104.5%,证明此传感器可以满足实际水样中痕量Hg2+的分析要求。

Specific Hg2+ Quantitation Using Intramolecular Split G-Quadruplex DNAzyme

G-quadruplex DNAzymes are peroxidase-like complexes formed by Hemin and some nucleic acid G-quadruplexes.Based on this kind of DNAzymes,several chemical sensors and biosensors have been developed.To improve the selectivity of G-quadruplex DNAzyme-based Hg2+ sensors,intramolecular split G-quadruplex DNAzyme was introduced in the design of Hg2+ sensor.For this new Hg2+ sensor,G-quadruplex-forming G-rich sequence was divided into two parts that were linked to the two ends of a Hg2+-sensing sequence.In the absence of Hg2+,G-quadruplex structure could not be formed because G-rich sequence was partly caged into an intermolecular duplex.In the presence of Hg2+,however,the stabilization of the T-T mismatch by Hg2+ could promote the folding of the Hg2+-sensing sequence into an intramolecular duplex,accompanied by the destruction of the intermolecular duplex and the formation of an intramolecular split G-quadruplex,which can form catalytically active G-quadruplex DNAzyme upon hemin binding.Utilizing the increase of peroxidase activity of the sensing system,a Hg2+ detection method has been developed.Using this method,Hg2+ quantitation can be achieved in the concentration ranges of 50-500 nmol/L and 2.0-7.5 μmol/L,with a detection limit of 47 nmol/L.The use of split G-quadruplex DNAzyme can increase the dependence of the sensor on Hg2+,thus,increase the selectivity of the sensor.The method was applied to the determination of Hg2+ in different water samples with the average recovery of 97.5%-104.5%.