Li Zhengtao, Lavergne Thomas, Malyshev Denis A, Zimmermann Jörg, Adhikary Ramkrishna, Dhami Kirandeep, Ordoukhanian Phillip, Sun Zhelin, Xiang Jie, Romesberg Floyd E
Department of Chemistry and Dr. P. Ordoukhanian Center for Protein and Nucleic Acid Research The Scripps Research Institute 10550 North Torrey Pines Road La Jolla, CA 92037.
Department of Electrical and Computer Engineering 9500 Gilman Drive University of California, San Diego La Jolla, CA 92093.
Chemistry. 2013 Oct 11;19(42):14205-14209. doi: 10.1002/chem.201302496. Epub 2013 Sep 11.
A class of replicable unnatural DNA base pairs formed between d5SICS and either dMMO2, dDMO, or dNaM were developed. To explore the use of these pairs to produce site-specifically labeled DNA, the synthesis of a variety of derivatives bearing propynyl groups, an analysis of their polymerase-mediated replication, and subsequent site-specific modification of the amplified DNA by Click chemistry is reported. With the d5SICS scaffold a propynyl ether linker is accommodated better than its aliphatic analogue, but not as well as the protected propargyl amine linker explored previously. It was also found that with the dMMO2 and dDMO analogues, the dMMO2 position para to the glycosidic linkage is best suited for linker attachment and that although aliphatic and ether-based linkers are similarly accommodated, the direct attachment of an ethynyl group to the nucleobase core is most well tolerated. To demonstrate the utility of these analogues, a variety of them were used to site-selectively attach a biotin tag to the amplified DNA. Finally, we use d5SICS(CO) -dNaM to couple one or two proteins to amplified DNA, with the double labeled product visualized by atomic force microscopy. The ability to encode the spatial relationships of arrayed molecules in PCR amplifiable DNA should have important applications, ranging from SELEX with functionalities not naturally present in DNA to the production, and perhaps "evolution" of nanomaterials.
开发了一类在d5SICS与dMMO2、dDMO或dNaM之间形成的可复制非天然DNA碱基对。为了探索利用这些碱基对产生位点特异性标记的DNA，本文报道了多种带有丙炔基的衍生物的合成、它们的聚合酶介导复制分析以及随后通过点击化学对扩增DNA进行的位点特异性修饰。对于d5SICS支架，丙炔基醚连接子比其脂肪族类似物更易容纳，但不如先前探索的受保护炔丙基胺连接子。还发现，对于dMMO2和dDMO类似物，糖苷键对位的dMMO2位置最适合连接子连接，并且尽管脂肪族和醚基连接子的容纳情况相似，但乙炔基直接连接到核碱基核心的耐受性最好。为了证明这些类似物的实用性，使用了多种类似物将生物素标签位点选择性地连接到扩增的DNA上。最后，我们使用d5SICS(CO)-dNaM将一种或两种蛋白质连接到扩增的DNA上，通过原子力显微镜观察到双标记产物。在可PCR扩增的DNA中编码阵列分子空间关系的能力应具有重要应用，范围从具有DNA中天然不存在的功能的SELEX到纳米材料的生产以及可能的“进化”。
