用于检测鸟嘌呤及其类似物的G四链体核酸探针的开发

Guanine is one of the basic units that make up the genetic material of the organism. It produces some analogues in the metabolic process of the organism, such as xanthine, 8-oxoguanine and uric acid. Guanine and its analogues are involved in many biological processes whose abnormalities can lead to a series of diseases, such as gout. Detection of guanine and its analogues can provide information for the prevention and diagnosis of many diseases. There were many studies on the detection methods of guanine and its analogues, but few of them were used in practical application. A good detection method requires sufficient sensitivity and specificity. The key difficulty was to find ligands that specifically bind to guanine or its analogues. Recently, SELEX technology had been widely used to screen aptamers for guanine and its analogues. In a latest study we found that a guanine-vacancy-bearing G-quadruplex may serve as a new class of aptamers. A guanine plane of such a G-quadruplex contained a gap which was specifically filled with a guanine base and formed four hydrogen bonds. In this project we wished to screen a class of guanine-vacancy-bearing G-quadruplex with high affinity for guanine. Using these G-quadruplexes as the basic skeleton, probes were designed to achieve sensitive and specific detection of guanine and its analogues.

鸟嘌呤是组成生物体遗传物质基本单元之一，其在生物体的代谢过程中会生成一些类似物，比如黄嘌呤，8氧鸟嘌呤，尿酸等。鸟嘌呤及其类似物参与许多生物学过程，它们的浓度异常会导致一系列疾病，比如痛风等。对鸟嘌呤及其类似物的检测可以为疾病的预防和诊断提供信息。有关鸟嘌呤及其类似物的检测方法的研究有很多，而实际推广应用的方法很少。一种好的检测方法需要有足够的灵敏度和特异性，其关键难点是寻找对鸟嘌呤或者其类似物特异性结合的配体，近年来SELEX技术被广泛用于筛选鸟嘌呤及其类似物的核酸适配体。本实验室最新研究发现的一种缺口G四链体可能成为一类新型核酸适配体。这种G四链体的一个鸟嘌呤平面上含有一个缺口，其能够特异性的被鸟嘌呤碱基填入并形成4个氢键。本项目中，我们希望筛选得到一类对鸟嘌呤有高亲和力的缺口G四链体，并以这种G四链体为基本骨架设计探针，实现对鸟嘌呤及其类似物的灵敏且特异的检测。

鸟嘌呤是组成生物体遗传物质的基本单元之一，对鸟嘌呤及其类似物的检测可以为疾病的预防和诊断提供有用的信息。针对鸟嘌呤及其类似物的检测方法的研究持续多年，但是实际推广应用的方法很少。.近年来的研究中，我们发现了一种能够特异性识别溶液中鸟嘌呤的缺口型G-四链体。这种G-四链体结构是一种非常有潜力的检测鸟嘌呤的核酸探针，它可以区分鸟嘌呤的不同代谢产物，唯一不足之处是结合鸟嘌呤的能力不高。为了开发具有应用价值的鸟嘌呤探针，我们首先筛选出一批能够结合鸟嘌呤及其类似物的缺口型G-四链体结构并对它们进行了鉴定；然后在缺口G-四链体的基础上加入一个结合鸟嘌呤平面的多肽，制备了一种多肽-G-四链体的复合型探针。当溶液中的鸟嘌呤填入缺口G-四链体的鸟嘌呤平面后，RHAU多肽能够进一步稳定鸟嘌呤平面，因此这种复合型探针相比单纯核酸探针结合鸟嘌呤的能力得到显著提升。该探针的后续开发还在进行。.在开发该复合型探针的时候，我们意外发现鸟嘌呤-RHAU多肽偶联的化合物能够选择性识别不同的核酸高级结构，比如，单个鸟嘌呤偶联的多肽能够特异性识别并稳定缺口型G-四链体，而三个鸟嘌呤偶联的多肽则能够特异性识别G-三链体。这些发现对靶向G-四链体的药物设计有极大的启示意义，相关成果已经发表。.此外我们还发现人基因组DNA上PDGFRB基因启动子区存在的一个缺口型G四链体，它能够响应环境鸟嘌呤的变化并且能够影响基因转录的水平。该缺口G-四链体是一种天然的鸟嘌呤探针，相关研究将在未来的工作中完成。