EFFECT OF P-CHIRALITY OF OLIGO(DEOXYRIBONUCLEOSIDE PHOSPHOROTHIOATE)SON THE ACTIVITY OF TERMINAL DEOXYRIBONUCLEOTIDYL TRANSFERASE

Phosphorothioate analogues of oligonucleotides (PS-oligos) of predeter mined chirality at the phosphorus atom at each internucleotide linkage have been used as primers for terminal deoxyribonucleotidyl transfera se (TdT, EC 2.7.7.31). The enzyme catalyzes efficient elongation of PS primers in which all phosphorothioate internucleotide linkages are un iformly of the [R-P] configuration, while the presence of the linkage( s) of the [S-P] configuration significantly decreases or completely in hibits the primer extension. Our results indicate that for the elongat ion of phosphorothioate oligomers the most important is the internucle otide bond located between the second and the third nucleoside from th e 3'-end. The presence of [S-P] linkage at this position strongly redu ces the enzyme activity while the [R-P] bond allo,vs for effective elo ngation of the primer. The activity of the enzyme is also influenced b y base composition and sequence of phosphorothioate primer as well as the dNTP used for elongation process. (C) 1998 Federation of European Biochemical Societies.