Identification and Characterization of Second-Generation Invader Locked Nucleic Acids (LNAs) for Mixed-Sequence Recognition of Double-Stranded DNA

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• 作者：Sujay P. Sau ; Andreas S. Madsen ; Peter Podbevsek ; Nicolai K. Andersen ; T. Santhosh Kumar ; Sanne Andersen ; Rie L. Rathje ; Brooke A. Anderson ; Dale C. Guenther ; Saswata Karmakar ; Pawan Kumar ; Janez Plavec ; Jesper Wengel ; Patrick J. Hrdlicka
• 刊名：Journal of Organic Chemistry
• 出版年：2013
• 出版时间：October 4, 2013
• 年：2013
• 卷：78
• 期：19
• 页码：9560-9570
• 全文大小：524K
• 年卷期：v.78,no.19(October 4, 2013)
• ISSN：1520-6904

文摘

The development of synthetic agents that recognize double-stranded DNA (dsDNA) is a long-standing goal that is inspired by the promise for tools that detect, regulate, and modify genes. Progress has been made with triplex-forming oligonucleotides, peptide nucleic acids, and polyamides, but substantial efforts are currently devoted to the development of alternative strategies that overcome the limitations observed with the classic approaches. In 2005, we introduced Invader locked nucleic acids (LNAs), i.e., double-stranded probes that are activated for mixed-sequence recognition of dsDNA through modification with 鈥?1 interstrand zippers鈥?of 2鈥?N-(pyren-1-yl)methyl-2鈥?amino-伪-l-LNA monomers. Despite promising preliminary results, progress has been slow because of the synthetic complexity of the building blocks. Here we describe a study that led to the identification of two simpler classes of Invader monomers. We compare the thermal denaturation characteristics of double-stranded probes featuring different interstrand zippers of pyrene-functionalized monomers based on 2鈥?amino-伪-l-LNA, 2鈥?N-methyl-2鈥?amino-DNA, and RNA scaffolds. Insights from fluorescence spectroscopy, molecular modeling, and NMR spectroscopy are used to elucidate the structural factors that govern probe activation. We demonstrate that probes with +1 zippers of 2鈥?O-(pyren-1-yl)methyl-RNA or 2鈥?N-methyl-2鈥?N-(pyren-1-yl)methyl-2鈥?amino-DNA monomers recognize DNA hairpins with similar efficiency as original Invader LNAs. Access to synthetically simple monomers will accelerate the use of Invader-mediated dsDNA recognition for applications in molecular biology and nucleic acid diagnostics.