Polynucleotide kinase from rat-liver nuclei. Purification and properties.
A polynucleotide kinase, which catalyzes the phosphorylation of 5'-hydroxyl ends of deoxyribonucleic acid in the presence of adenosine triphosphate, has been purified 260-fold with a yield of 14% from 0.15 M NaCl extracts of rat liver nuclei. The purified enzyme has a pH optimum of 5.5. The enzyme is reversible inhibited by p-chloromercuribenzoate. The S0.5 value (ligand concentration required for a half-maximal activity) for ATP is 2.5 muM. A bivalent cation is essential for the reaction and S0.5 values for Mg2+, Ca2+ and Mn2+ are 3.3 mM, 4 mM and 0.05 mM respectively. Pyrophosphate remarkable inhibits the activity with I0.5 value (ligand concentration required for a half-maximal inhibition) of 0.2 mM, and sulfate, with I0.5 of 0.5 mM, whereas phosphate weakly inhibits the activity with I0.5 of about 20 mM. An apparent molecular weight of the purified enzyme is estimated to be 8 X 10(4) by gel filtration on a column of Sephadex G-150, and the Stokes radius of the enzyme molecule is shown to be about 0.36 nm. Sucrose density gradient centrifugation reveals that the enzyme has a sedimentation coefficient of about 4.4 S.
D000255:Adenosine Triphosphate / Q000494:pharmacology; D000818:Animals; D002118:Calcium / Q000494:pharmacology; D002467:Cell Nucleus / Q000201:enzymology; D002729:Chloromercuribenzoates / Q000494:pharmacology; D004247:DNA / Q000378:metabolism; D011756:Diphosphates / Q000494:pharmacology; D006863:Hydrogen-Ion Concentration; D008099:Liver / Q000201:enzymology* / Q000648:ultrastructure; D008274:Magnesium / Q000494:pharmacology; D008297:Male; D008345:Manganese / Q000494:pharmacology; D008970:Molecular Weight; D010710:Phosphates / Q000378:metabolism; D010770:Phosphotransferases / Q000378:metabolism*; D011116:Polynucleotide 5'-Hydroxyl-Kinase / Q000302:isolation & purification / Q000378:metabolism*; D051381:Rats; D013431:Sulfates / Q000494:pharmacology
D002729:Chloromercuribenzoates; D011756:Diphosphates; D010710:Phosphates; D013431:Sulfates; D008345:Manganese; D000255:Adenosine Triphosphate; D004247:DNA; D010770:Phosphotransferases; D011116:Polynucleotide 5'-Hydroxyl-Kinase; D008274:Magnesium; D002118:Calcium