The Polymerization of Guanosine Diphosphate by Polynucleotide Phosphorylase

Polynucleotide phosphorylase catalyzes the reversible polymerization of nucleoside diphosphates (1). With enzyme preparations from Escherichia coli (2), Azotobacter agile (3), and Micrococcus lysodeikticus (4, 5) a reaction has been observed with single additions of adenosine, uridine, cytosine, or inosine diphosphate to form the corresponding homopolymer. However, the polymerization of guanosine diphosphate represents a special case. Littauer and Kornberg (2) observed no polymerization reaction upon incubating GDP w-ith fractions from E. coZi. In experiments with polynucleotide phosphorylase from A. agile, Grunberg-Manago et al. (3) noted a very slow release of Pi from GDP, but this reaction stopped far short of the equilibrium point found for ADP and UDP. In contrast, when mixed with other nucleoside diphosphates, GDP is well utilized and polymers such as poly AGUCl are formed (3). The experiments described below show that GDP, when present alone, is not polymerized even after many hours by enzyme fractions from A. agile or E. coli. However, polymerization of GDP does take place in the presence of oligonucleotides such as pApApA and ApApU, and guanosine monophosphate units are added to the primer. No reaction occurs with GDP and oligonucleotides, such as ApApUp, that do not contain an unsubstituted hydroxyl group at carbon 3’ of the terminal nucleoside residue. Highly purified fractions of A. agile polynucleotide phosphorylase, kindly supplied by Drs. Mii and Ochoa (B), catalyze the polymerization of ADP, UDP, CDP, and IDP only after a lag period. This lag can be overcome by ribonucleic acid and certain other polymers (6)) as well as by oligoribonucleotides of various types, including pApApA, ApApU and ApApUp (see preceding paper (7)). Accordingly, GDP differs from other nucleoside diphosphates in that its polymerization requires the presence of oligonucleotides of the type that can be incorporated into the polymer.