Murakami Kazuyoshi, Zhao Jing, Yamasaki Kazuhiko, Miyagishi Makoto
Molecular Composite Medicine Research Group, Biomedical Research Institute, National Institute of Advanced Industrial Science and Technology, Tsukuba-shi, Ibaraki 305-8566, Japan.
Biomed Rep. 2017 Jun;6(6):615-626. doi: 10.3892/br.2017.899. Epub 2017 May 3.
Extracellular vesicles are particles in mammalian body fluids that have attracted considerable attention as biomarkers for various diseases. In the present study, the authors isolated RNA aptamers with an affinity for extracellular vesicles from two library pools that encoded randomized sequences of different lengths. After the several rounds of selection, two conserved motifs are identified in the sequences that are obtained by next-generation sequencing. Most of the sequences were predicted to adopt a secondary structure that consisted of a non-conserved stem structure and a conserved loop sequence. Two minimal similar sequences are synthesized and confirmed the ability of these sequences to bind to extracellular vesicles. Circular dichroism spectroscopy and melting temperature analysis demonstrated that the aptamers were able to form a G-quadruplex structure in their loop regions and these structures were stabilized by potassium ions. Consistent with these structural data, the affinity of each aptamer for extracellular vesicles was dependent on potassium ions. The aptamers that were identified may be useful molecular tools for the development of diagnostic methods that utilize body fluids, such as blood, saliva and urine.
细胞外囊泡是哺乳动物体液中的颗粒，作为各种疾病的生物标志物已引起了广泛关注。在本研究中，作者从两个编码不同长度随机序列的文库池中分离出对细胞外囊泡具有亲和力的RNA适体。经过几轮筛选，通过下一代测序获得的序列中鉴定出两个保守基序。大多数序列预计会形成一种二级结构，该结构由一个非保守的茎结构和一个保守的环序列组成。合成了两个最小相似序列，并证实了这些序列与细胞外囊泡结合的能力。圆二色光谱和熔解温度分析表明，适体能够在其环区域形成G-四链体结构，并且这些结构由钾离子稳定。与这些结构数据一致，每个适体对细胞外囊泡的亲和力取决于钾离子。所鉴定的适体可能是开发利用血液、唾液和尿液等体液的诊断方法的有用分子工具。
