Williams D M, Benseler F, Eckstein F
Abteilung Chemie, Max-Planck-Institut für Experimentelle Medizin, Göttingen, FRG.
Biochemistry. 1991 Apr 23;30(16):4001-9. doi: 10.1021/bi00230a027.
2'-Fluorothymidine (Tf) was synthesized via an improved procedure with (diethylamino)sulfur trifluoride. The compatibility of the analogue with DNA synthesis via the phosphoramidite method was demonstrated after complete enzymatic digestion of the oligonucleotides d(Tf11T) and d(Tf3T), the sole products of which were 2'-fluorothymidine and thymidine in the expected ratio. The 2'-fluorothymidine was also incorporated into the EcoRV recognition sequence (underlined), within the complementary oligonucleotides d(CAAACCGATATCGTTGTG) and d(CACAACGATATCGGTTTG). Thermal melting characteristics of these duplexes showed a significant decrease in stability only when both of the thymidine residues in one of the strands were replaced. In contrast, when all of one strand of a duplex contained 2'-fluorothymidine, as in d(Tf11T).d(A12), a substantially higher Tm and cooperativity of melting was observed relative to the unmodified structure. EcoRV cleaved a duplex that contained a 2'-fluorothymidine at the scissile linkage in each strand at two-thirds of the rate obtained for the unmodified structure. A duplex containing two 2'-fluorothymidine residues in one strand and none in the other was cleaved at one-third of the rate in its unsubstituted strand, whereas the cleavage rate was reduced to 22% in its modified strand.
2'-氟胸苷（Tf）通过一种改进的方法，利用三氟（二乙氨基）硫合成。在对寡核苷酸d（Tf11T）和d（Tf3T）进行完全酶切后，证明了该类似物与通过亚磷酰胺法进行DNA合成的兼容性，其唯一产物是预期比例的2'-氟胸苷和胸苷。2'-氟胸苷也被掺入到互补寡核苷酸d（CAAACCGATATCGTTGTG）和d（CACAACGATATCGGTTTG）中的EcoRV识别序列（下划线部分）内。这些双链体的热解链特性表明，只有当一条链中的两个胸苷残基都被取代时，稳定性才会显著降低。相比之下，当双链体的一条链全部包含2'-氟胸苷时，如d（Tf11T）.d（A12），相对于未修饰的结构，观察到解链温度（Tm）显著更高且解链协同性更高。EcoRV切割每条链在可切割位点含有2'-氟胸苷的双链体的速率是未修饰结构的三分之二。一条链中含有两个2'-氟胸苷残基而另一条链中没有的双链体，其未取代链的切割速率为未修饰时的三分之一，而其修饰链的切割速率降至22%。
