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Monofunctional Substrates of Polynucleotide Phosphorylase
The Monoaddition of 2′(3′)‐O‐Isovaleryl‐nucleoside Diphosphate to an Initiator Oligonucleotide
Definition. A260 unit, the quantity of material contained in 1 ml of a solution which has an absorbance of 1 at 260 nm, when measured in a 1‐cm pathlength cell.
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.
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