The enzymatic phosphorylation of ribonucleic acid and deoxyribonucleic acid. II. Further properties of the 5'-hydroxyl polynucleotide kinase.

The specificity of the enzyme purified from TZ-infected Escherichia coli which catalyzes the phosphorylation of 5’hydroxyl ends of ribonucleic acid and deoxyribonucleic acid has been examined. The enzyme also catalyzes the phosphorylation of 5’-hydroxyl groups of polynucleotides as well as 3’-mononucleotides. There is no detectable phosphorylation of nucleosides and adenosine 2’-phosphate is virtually inactive as a phosphate acceptor. The enzyme is also capable of using guanosine, @dine, and uridine trlphosphates as well as adenosine triphosphate as the phosphorylating agent. The enzyme catalyzes the quantitative phosphorylation of available S-hydroxyl groups occurring in a variety of different DNA preparations. When used in conjunction with alkaline phosphatase, the enzymatic phosphorylation of DNA measures the total amount of S’-hydroxyl and 5’-phosphate ends present in DNA. The 5’-hydroxyl polynucleotide kinase was used to measure the amount of 5’-hydroxyl ends formed in DNA degraded by sonic oscillation and alkali and heat denaturation. In all of the cases, virtually no additional Shydroxyl groups were formed. The 5’-hydroxyl kinase has also been detected in nuclei of rat liver and the activity partially characterized.