Polynucleotide Phosphorylase

Polynucleotide phosphorylase (PNPase) is the first enzyme that can catalyze the formation of polyribonucleotides with a 3′,5′-phosphodiester bond. In the forward reaction, long polyribonucleotides are synthesized from various ribonucleoside diphosphates, with elimination of inorganic orthophosphate. Each of the four common ribonucleoside diphosphates can serve separately as a substrate for the polymerization reaction, leading to the formation of homopolymers. Polymerization of a mixture of nucleoside diphosphates that contain different bases results in the formation of a random copolymer. In the reverse reaction, the enzyme catalyzes the breakdown of polyribonucleotides by phosphorolytic cleavage of the internucleotide bonds. PNPase also catalyzes an exchange reaction between ³²P-labeled inorganic phosphate and the β-phosphate of nucleoside diphosphates. This chapter summarizes the current knowledge regarding the PNPase enzyme, its purification, properties, and the various reactions it catalyzes. It emphasizes the wide range of research applications that are in use with PNPase enzyme. PNPase is a useful tool for the synthesis of polynucleotides with varied composition, both in the presence and in the absence of primer oligonucleotides.