Fundamental Genetic Processes and Mechanisms

Photoreactive derivatives of oligonucleotides containing arylazido groups at 5’-phosphate introduced by chemical synthesis extensively are used for the investigation of the components of DNA replication/repair systems by the methods of photoaffinity modification. We proposed that ATP derivatives bearing photoreactive substitutions at γ-phosphate might turn out substrates for T4 polynucleotide kinase (T4 PNK) and this allows introducing different groups into the 5’-OH end of oligonucleotides. A set of γphosphoamides of ATP was synthesized and shown to be substrates of this enzyme. Photoreactive oligonucleotide conjugate obtained by using of the γ-N[-(4-azido-2,3,5,6-tetrafluorobenzoyl)-aminoethyl] amide ATP and 17-mer deoxyribooligonucleotide as substrates has been used for the photoaffinity labelling of human replication protein A (RPA) and flap endonuclease 1 (FEN 1). It can be proved that tested set of the γ-amides of ATP can be applied for the realization of the new enzymatic approach synthesis of 5'-end-phosphate substituted derivatives of riboand deoxyribooligonucleotides.