Rationally manipulating aptamer binding affinities in a stem-loop molecular beacon.
Rationally manipulating aptamer binding affinities in a stem-loop molecular beacon.
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DOI：
10.1021/bc500286r
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发表时间：
2014-10-15
影响因子：
4.7
通讯作者：
Strouse, Geoffrey F.
中科院分区：
文献类型：
--
作者：
Armstrong, Rachel E.;Strouse, Geoffrey F.
对靶配体具有高度特异性的单链DNA序列称为适体。虽然将适体序列并入茎-环分子信标中已成为光学生物传感器中的重要工具，但确定结合亲和力的大小及其与适体序列在茎-环结构中的放置的关系的设计原理尚未明确定义。通过将适体沿着分子信标的环区域受控放置，观察到对于Huizenga和Szostak ATP DNA适体序列，结合亲和力可调节超过4个数量级（1.3 nM - 203 μM）。观察到，对于完全暴露的序列，Kd增强，当适体是信标的茎区的一部分时，结合亲和力降低。ΔG值的分析表明茎中的适体杂交长度与其观察到的Kd之间存在明显的相关性。使用纳米金属表面能量转移探针方法监测ATP与适体序列的结合允许观察两个ATP结合事件之间的负协同性。维持这种ATP适体的高结合亲和力和观察ATP结合的两个单独的Kd，表明NSET作为一种有效的，非操纵性的，光学方法用于跟踪生物分子的变化。
Single-stranded DNA sequences that are highly specific for a target ligand are called aptamers. While the incorporation of aptamer sequences into stem-loop molecular beacons has become an essential tool in optical biosensors, the design principles that determine the magnitude of binding affinity and its relationship to placement of the aptamer sequence in the stem-loop architecture are not well defined. By controlled placement of the aptamer along the loop region of the molecular beacon, it is observed that the binding affinity can be tuned over 4 orders of magnitude (1.3 nM – 203 μM) for the Huizenga and Szostak ATP DNA aptamer sequence. It is observed that the Kd is enhanced for the fully exposed sequence, with reduced binding affinity when the aptamer is part of the stem region of the beacon. Analysis of the ΔG values indicate a clear correlation between the aptamer hybridized length in the stem and its observed Kd. The use of a nanometal surface energy transfer probe method for monitoring ATP binding to the aptamer sequence allows the observation of negative cooperativity between the two ATP binding events. Maintenance of the high binding affinity of this ATP aptamer and the observation of two separate Kd’s for ATP binding indicate NSET as an effective, nonmanipulative, optical method for tracking biomolecular changes.
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影响因子：
17.1
作者：
Lu, Wentong;Arumugam, Ranjini;Senapati, Dulal;Singh, Anant K.;Arbneshi, Tahir;Khan, Sadia Afrin;Yu, Hongtao;Ray, Paresh Chandra
通讯作者：
Ray, Paresh Chandra
影响因子：
--
作者：
Lin, CH;Patel, DJ
通讯作者：
Patel, DJ
影响因子：
12.6
作者：
Ikebukuro, K;Kiyohara, C;Sode, K
通讯作者：
Sode, K
影响因子：
14.8
作者：
Haller, Andrea;Rieder, Ulrike;Micura, Ronald
通讯作者：
Micura, Ronald
影响因子：
3.7
作者：
Breshike, Christopher J.;Riskowski, Ryan A.;Strouse, Geoffrey F.
通讯作者：
Strouse, Geoffrey F.