Aptameric enzyme subunit for biosensing based on enzymatic activity measurement
Aptameric enzyme subunit for biosensing based on enzymatic activity measurement
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DOI：
10.1021/ac060254o
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发表时间：
2006-05-15
影响因子：
7.4
通讯作者：
Ikebukuro, Kazunori
中科院分区：
文献类型：
--
作者：
Yoshida, Wataru;Sode, Koji;Ikebukuro, Kazunori
核酸适体酶亚基（aptamericenzyme subunit，AES）是由酶抑制性适体和靶分子结合性适体组成的DNA适体，是一种用于靶分子生物传感的分子。我们使用凝血酶抑制适体作为抑制酶活性的适体。凝血酶抑制适体折叠成G-四联体结构，这在其抑制活性中起重要作用。作为靶分子结合适体，将腺苷结合适体插入凝血酶抑制适体的G-四联体结构中，以使G-四联体结构在腺苷识别时发生变化。在本研究中，G-四重结构的变化导致凝血酶抑制活性的变化，并成功地检测到腺苷通过测量在均匀的溶液中的凝血酶活性没有结合/自由分离。我们构建了两种AES，其中一种结构是通用的，可以用于设计任何靶分子结合适体。由于酶活性是测量的，AES能够在均相测定中简单且高灵敏度地检测靶分子。
The aptameric enzyme subunit (AES), which is a DNA aptamer composed of an enzyme-inhibiting aptamer and a target molecule-binding aptamer, has been developed for the biosensing of target molecules. We used a thrombin-inhibiting aptamer as the aptamer that inhibits enzymatic activity. The thrombin-inhibiting aptamer folds into the G-quartet structure, which plays an important role in its inhibitory activity. As a target molecule-binding aptamer, an adenosine-binding aptamer was inserted into the G-quartet structure of the thrombin-inhibiting aptamer to enable the change of the G-quartet structure upon the recognition of adenosine. In the present study, the change in the G-quartet structure led to a change in the thrombin inhibition activity, and adenosine was successfully detected by measuring the thrombin activity in a homogeneous solution without bound/free separation. We constructed two kinds of AESs; one of the structures is universal and can be used for designing any target molecule-binding aptamer. Since the enzyme activity is measured, AESs enable the simple and high-sensitivity detection of target molecules in a homogeneous assay.