Ribonucleotide reductase, an enzyme found in all prokaryotic and eukaryotic cells that synthesize DNA, is induced by herpes simplex virus (HSV). In this study the effect of anti-HSV antiserum on the induced ribonucleotide reductase has been examined and the ability of different temperature-sensitive (ts) mutants of HSV-1 to induce the enzyme has been investigated. The HSV-1-induced ribonucleotide reductase was inhibited by antiserum raised against infected cell lysates but not by preimmune serum. The wild-type (ts+) virus induced similar levels of ribonucleotide reductase at 31 degrees C and 38.5 degrees C (the permissive and non-permissive temperatures respectively for the ts mutants). All ts mutants induced approximately wild-type levels of the enzyme at 31 degrees C. At 38.5 degrees C, two of the four ts mutants studied also induced wild-type levels of enzyme but ts G failed to induce any activity while ts K induced variable but low levels. The enzyme activity induced by ts G at 31 degrees C was thermolabile both in vivo and in vitro. These results provide the first strong evidence that the induced ribonucleotide reductase activity is at least partially virus-coded.