Sterical recognition by T4 polynucleotide kinase of non-nucleosidic moieties 5′-attached to oligonucleotides

The ability of T4 polynucleotide kinase (PNK) to phosphorylate non-nucleosidic moieties 5′-attached to oligodeoxynucleotides (ODNs) has been investigated. Non-nucleosidic phosphoramidite units were prepared from ethane-1,2-diol and propane-1,3-diol backbones. Some of them corresponded to pure enantiomers. They were used to obtain the corresponding 5′-end modified oligothymidylates X(pdT)10. The free primary hydroxyl of the non-nucleosidic moieties (X) of these oligomers was phosphorylated by PNK. We report the stereo-selective phosphorylation of the L form of the 5′-end attached non-nucleosidic chiral fragments; the non-chiral moieties were completely phosphorylated. Dimers of glycerol analogue and thymidine 3′-phosphate were not recognized by PNK and the shortest modified ODN able to be phosphorylated was a trinucleotide X(pdT)3. A modified X(pdT)10, bearing a cyclic abasic site (X) at its 5′-end, was prepared by chemical synthesis from 1,2-dideoxyribose phosphoramidite and was phosphorylated with a 90% yield.