Incorporating 2-thiouracil into short dsRNA-binding PNAs for enhanced recognition of A-U pairs and for targeting a microRNA hairpin precursor.
Incorporating 2-thiouracil into short dsRNA-binding PNAs for enhanced recognition of A-U pairs and for targeting a microRNA hairpin precursor.
复制标题
将 2-硫氧嘧啶掺入短 dsRNA 结合 PNA 中，以增强对 A-U 对的识别并靶向 microRNA 发夹前体。
作者：
A. Ong;Desiree;Manchugondanahalli S. Krishna;Kiran M. Patil;K. Okamura;Gang Chen
化学修饰的短肽核酸（PNAs）在接近生理条件下通过形成大沟PNA·RNA-RNA三链体识别RNA双链体，在开发RNA靶向探针和治疗药物方面显示出巨大的潜力。胸腺嘧啶（T）和尿嘧啶（U）通常被掺入PNA中，通过形成大沟T·A-U和U·A-U三联体来识别A-U对。将修饰的核碱基2-硫氧嘧啶（s2 U）掺入三链体形成寡核苷酸（TFO）中可稳定DNA和RNA三链体。尿嘧啶的巯基化导致三链体形成的脱水能量损失降低以及N3氮原子的pKa降低，这可能导致除了增强的碱基堆积相互作用之外的氢键结合改善，类似于先前报道的假异胞嘧啶的巯基化效应（J至L取代）。在这里，我们将s2 U短PNA，随后结合一系列的s2 U修饰的PNA的研究。我们通过非变性聚丙烯酰胺凝胶电泳（PAGE）和热熔解实验证明，s2 U和L掺入到dsRNA结合的PNA（dbPNAs）中，分别以位置无关的方式增强了RNA双链体中A-U和G-C对的识别，而与DNA双链体没有明显的结合。在dbPNA中组合s2 U和L修饰有利于增强对dsRNA的识别，并保持与dsRNA的选择性结合超过ssRNA。我们通过无细胞测定进一步证明了s2 U-和L-修饰的dbPNA在以底物特异性方式抑制前微RNA-198成熟中的应用。因此，s2 U修饰的dbPNA通常可用于增强和选择性识别RNA双链体和调节RNA功能。
Chemically-modified short Peptide Nucleic Acids (PNAs) recognize RNA duplexes at near physiological conditions by major-groove PNA·RNA-RNA triplex formation and show a great promise in developing RNA-targeting probes and therapeutics. Thymine (T) and uracil (U) are often incorporated into PNAs to recognize A-U pairs through major-groove T·A-U and U·A-U triple formation. Incorporation of a modified nucleobase, 2-thiouracil (s2U), into triplex-forming oligonucleotides (TFOs) stabilizes both DNA and RNA triplexes. Thiolation of uracil causes a decrease of the dehydration energy penalty for triplex formation as well as a decrease of the pKa of N3 nitrogen atom, which may result in improved hydrogen bonding in addition to enhanced base stacking interactions, similar to the previously reported thiolation effect of pseudoisocytosine (J to L substitution). Here, we incorporated s2U into short PNAs, followed by binding studies of a series of s2U-modified PNAs. We demonstrated by nondenaturing polyacrylamide gel electrophoresis (PAGE) and thermal melting experiments that s2U and L incorporated into dsRNA-binding PNAs (dbPNAs) enhance the recognition of A-U and G-C pairs, respectively, in RNA duplexes in a position-independent manner, with no appreciable binding to DNA duplex. Combining s2U and L modifications in dbPNAs facilitates enhanced recognition of dsRNAs and maintains selective binding to dsRNAs over ssRNAs. We further demonstrated through a cell-free assay the application of the s2U- and L-modified dbPNAs in the inhibition of the pre-microRNA-198 maturation in a substrate specific manner. Thus, s2U-modified dbPNAs may be generally useful for the enhanced and selective recognition of RNA duplexes and for the regulation of RNA functions.
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影响因子：
15
作者：
Li, Ming;Zengeya, Thomas;Rozners, Eriks
通讯作者：
Rozners, Eriks
影响因子：
4
作者：
Muse, Oluwatoyosi;Zengeya, Thomas;Mwaura, Juddy;Hnedzko, Dziyana;McGee, Dennis W.;Grewer, Christof T.;Rozners, Eriks
通讯作者：
Rozners, Eriks
通过含有新型碱基类似物的三链体形成寡核苷酸靶向哺乳动物细胞中中断的多嘌呤：多嘧啶序列。
影响因子：
2.9
作者：
Semenyuk,A;Darian,E;Liu,J;Majumdar,A;Cuenoud,B;Miller,PS;MackerellJr,AD;Seidman,MM
通讯作者：
Seidman,MM
影响因子：
2.9
作者：
Carlucci, Marta;Kierzek, Elzbieta;Olejnik, Anna;Turner, Douglas H.;Kierzek, Ryszard
通讯作者：
Kierzek, Ryszard
影响因子：
12.4
作者：
Wilson RC;Doudna JA
通讯作者：
Doudna JA