SUMMARY The relative activity toward different nucleoside diphosphates remains essentially unchanged after 500-fold purification of polynucleotide phosphorylase of Azotobacter v&elan&i suggesting that a single enzyme is involved. The purified enzyme, virtually free of nuclease, contains small amounts of an oligoribonucleotide (with adenine, guanine, uracil, and cytosine in similar ratios as in whole cell Azotobacter RNA) which is released on denaturation of the protein. Although we do not know whether this oligonu- cleotide is a prosthetic group or a tightly clinging contaminant, it may act as a primer polynucleotide synthesis by the enzyme. No primer addition is required for the synthesis of polyadenylic acid when relatively large amounts of enzyme or Mg++ are used, but a requirement for added primer becomes apparent with small amounts of either component. Synthesis of polymers containing guanylic acid is always markedly dependent on addition of primer and requires much more enzyme. Under optimal con- ditions, the turnover number for polyadenylic acid synthesis (0.02