On the defect of a dCMP hydroxymethylase mutant of bacteriophage T4 showing enzyme activity in extracts

Infection by $\text{L}\text{13}$, a temperature-sensitive mutant of gene 42 of phage T4, the structural gene for dCMP hydroxymethylase, previously was shown not to form T4 DNA at nonpermissive temperatures. Yet the enzyme activity was found in extracts. Since inactivation of the enzyme was not reversible, we have examined acid-soluble extracts of cells infected at nonpermissive temperature by $\text{tsL}\text{13}$ for 5-hydroxymethyldCMP in order to determine whether the enzyme functioned $\text{in vivo}$. A double mutant of $\text{tsL13}$ and $\text{amB}\text{24}$ (5-hydroxymethyldCMP kinase) did not form the nucleotide at nonpermissive temperature, but the control, $\text{amB}\text{24}$, formed large quantities. From these results and previous temperature-shift studies it is suggested that the enzyme is normally activated to function $\text{in vivo}$ between 5 and 8 minutes after infection.