STERICAL RECOGNITION BY T-4 POLYNUCLEOTIDE KINASE OF NON-NUCLEOSIDIC MOIETIES 5'-ATTACHED TO OLIGONUCLEOTIDES

The ability of T-4 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 stereoselective phosphorylation of the L form of the 5'-end attached non-nucleosidic chiral fragments; the nonchiral 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.