Monofunctional Substrates of Polynucleotide Phosphorylase

Abstract

Polynucleotide phosphorylase from Escherichia coli cells catalyzes the transfer of a single nucleotidyl residue from each of the 2′(3′)‐ O ‐isovalerylesters of ADP, GDP, CDP and UDP to the initiator oligonucleotide, A‐A‐A. The products of these reactions are identified as the 2′(3′)‐ O ‐isovaleryl esters of the corresponding tetranucleotides A‐A‐A‐A, A‐A‐A‐G, A‐A‐A‐C and A‐A‐A‐U. The 2′(3′)‐ O ‐isovaleryl residue can be removed from the product by treatment with aqueous methanolic ammonia under conditions that do not significantly damage the oligonucleotide chains. The monoaddition of 2′(3′)‐ O ‐acyl esters of nucleoside diphosphates to an oligonucleotide acceptor is proposed as a method for the stepwise synthesis of oligoribonucleotides of defined sequence.