Liebmann Meike, Di Pasquale Francesca, Marx Andreas
Fachbereich Chemie, Universität Konstanz, Universitätsstrasse 10, 78457 Konstanz, Germany.
Chembiochem. 2006 Dec;7(12):1965-9. doi: 10.1002/cbic.200600333.
The cross-linking of target proteins or nucleic acids to light-activatable ligands is an important tool for elucidating molecular interactions. Through the use of photoaffinity-labeling reagents, several new insights into nucleic acid interactions have been obtained, for example in DNA replication and repair. In most known photoprobes, the applied light-sensitive functionalities are placed directly at the nucleobase or are attached via linkers to either the nucleobase or the phosphate backbone. Here we describe the first photoprobe that bears a light-sensitive aryl(trifluoromethyl)diazirine at the sugar moiety of a DNA oligonucleotide. We devised a route for the synthesis of the modified nucleoside and its incorporation into an oligonucleotide. The photoactive species was proven to be stable under the conditions employed in routine automated DNA synthesis. The modified oligonucleotide was shown by subsequent photolabeling studies of human DNA polymerase beta to form a covalent complex to the enzyme upon irradiation with near-UV light.
将目标蛋白质或核酸与光可激活配体交联是阐明分子相互作用的重要工具。通过使用光亲和标记试剂，已经获得了一些关于核酸相互作用的新见解，例如在DNA复制和修复方面。在大多数已知的光探针中，所应用的光敏感官能团直接位于核碱基上，或者通过连接子连接到核碱基或磷酸骨架上。在这里，我们描述了第一种在DNA寡核苷酸的糖部分带有光敏感芳基（三氟甲基）二氮杂环丙烷的光探针。我们设计了一种合成修饰核苷并将其掺入寡核苷酸的路线。经证明，光活性物质在常规自动化DNA合成所用的条件下是稳定的。通过随后对人DNA聚合酶β的光标记研究表明，修饰的寡核苷酸在近紫外光照射下与该酶形成共价复合物。
