Isolation and characterization of two mutant forms of T4 polynucleotide kinase.
The purification of polynucleotide kinase from Escherichia coli infected by two different mutants in the T4 polynucleotide kinase (pseT) gene is described. The pseT 1 enzyme has virtually no 3' specific phosphatase activity and normal polynucleotide kinase activity. The pseT 47 enzyme has very little phosphatase activity and no kinase activity. However, enzyme isolated from a pseT 1, pseT 47 mixed infection appears to contain heterodimers with considerably more phosphatase activity. Thus, the pseT 47 mutation partially inactivates the phosphatase and totally inactivates the kinase. A study of the action of polynucleotide kinase on plasmid DNAs nicked to give a 3'-phosphate and a 5'-hydroxyl indicates that although the enzyme can catalyze both the removal of the 3'-phosphate and the insertion of a 5'-phosphate, there is no evidence for a concerted reaction involving both activities on the same polypeptide chain.
D000596:Amino Acids / Q000032:analysis; D004926:Escherichia coli / Q000201:enzymology*; D007700:Kinetics; D009154:Mutation; D010770:Phosphotransferases / Q000302:isolation & purification*; D011116:Polynucleotide 5'-Hydroxyl-Kinase / Q000235:genetics / Q000302:isolation & purification* / Q000378:metabolism; D013604:T-Phages / Q000201:enzymology* / Q000235:genetics
D000596:Amino Acids; D010770:Phosphotransferases; D011116:Polynucleotide 5'-Hydroxyl-Kinase