In vitro selection of aptamers with affinity for neuropeptide Y using capillary electrophoresis
In vitro selection of aptamers with affinity for neuropeptide Y using capillary electrophoresis
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DOI：
10.1021/ja052406n
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发表时间：
2005-07-06
影响因子：
15
通讯作者：
Bowser, MT
中科院分区：
文献类型：
--
作者：
Mendonsa, SD;Bowser, MT
采用毛细管电泳-指数富集配体系统进化法（CE-SELEX）筛选神经肽Y（NPY）的适体。这是CE-SELEX选择结合比自身小的靶分子的适体的第一个例子。CE-SELEX的局限性之一是，当适体结合靶标以促进级分收集时，适体必须表现出显著的迁移率变化。在这项研究之前，尚不清楚较小的靶标是否能够诱导足够大的移动性变化，以使CE-SELEX成功。NPY是一种36个氨基酸的肽（MW = 4272 g/mol），比选择中使用的80个碱基的ssDNA（约25 kDa）小得多。仅经过4轮选择就获得了解离常数为300 - 1000 nM的NPY结合适体。使用人胰腺多肽（hPP）测试适体的特异性。hPP是由36个氨基酸组成的多肽，与NPY的同源性约为50%。观察到对NPY的选择性高达hPP的42倍的适体。
Capillary electrophoresis-systematic evolution of ligands by exponential enrichment (CE-SELEX) was used to select aptamers for neuropeptide Y (NPY). This is the first example of a CE-SELEX selection for aptamers that bind a target molecule smaller than itself. One of the limitations of CE-SELEX is that the aptamer must exhibit a significant mobility shift when it binds the target to facilitate fraction collection. Before this study, it was not clear if smaller targets would be capable of inducing a large enough shift in mobility for CE-SELEX to be successful. NPY is a 36-amino acid peptide (MW = 4272 g/mol), much smaller than the 80-base ssDNA used in the selection (∼25 kDa). NPY binding aptamers with 300−1000 nM dissociation constants were obtained after only four rounds of selection. The specificity of the aptamers was tested using human pancreatic polypeptide (hPP). hPP is a 36-amino acid peptide with ∼50% homology with NPY. Aptamers with up to 42-fold selectivity for NPY over hPP were observed.