Department of Chemistry, University of Zürich, Winterthurerstrasse 190, 8057, Zürich, Switzerland.
Bioorganic Research Institute, Suntory Foundation for Life Sciences, 8-1-1 Seikadai, Seikacho, Soraku, 619-0284, Kyoto, Japan.
Chembiochem. 2018 Sep 17;19(18):1939-1943. doi: 10.1002/cbic.201800351. Epub 2018 Aug 8.
Metabolic incorporation of bioorthogonal functional groups into cellular nucleic acids can be impeded by insufficient phosphorylation of nucleosides. Previous studies found that 5azidomethyl-2'-deoxyuridine (AmdU) was incorporated into the DNA of HeLa cells expressing a low-fidelity thymidine kinase, but not by wild-type HeLa cells. Here we report that membrane-permeable phosphotriester derivatives of AmdU can exhibit enhanced incorporation into the DNA of wild-type cells and animals. AmdU monophosphate derivatives bearing either 5'-bispivaloyloxymethyl (POM), 5'-bis-(4-acetoxybenzyl) (AB), or "Protide" protective groups were used to mask the phosphate group of AmdU prior to its entry into cells. The POM derivative "POM-AmdU" exhibited better chemical stability, greater metabolic incorporation efficiency, and lower toxicity than "AB-AmdU". Remarkably, the addition of POM-AmdU to the water of zebrafish larvae enabled the biosynthesis of azide-modified DNA throughout the body.
生物正交功能基团在细胞核酸中的代谢掺入可能会受到核苷磷酸化不足的阻碍。先前的研究发现，在表达低保真胸苷激酶的 HeLa 细胞中，5-叠氮甲基-2'-脱氧尿苷（AmdU）被掺入 DNA 中，但野生型 HeLa 细胞则不行。在这里，我们报告说，AmdU 的可透过细胞膜的膦酸三酯衍生物可以增强掺入野生型细胞和动物的 DNA 中。AmdU 单磷酸酯衍生物带有 5'-双(异戊酰氧甲基）（POM）、5'-双-(4-乙酰氧基苄基）（AB）或“Protide”保护基团，用于在 AmdU 进入细胞之前对其磷酸基团进行掩蔽。POM 衍生物“POM-AmdU”比“AB-AmdU”具有更好的化学稳定性、更高的代谢掺入效率和更低的毒性。值得注意的是，将 POM-AmdU 添加到斑马鱼幼虫的水中，可使整个身体合成叠氮修饰的 DNA。
