Purification and properties of a DNA ligase from sea urchin embryos.
DNA ligase was purified about 2,000-fold from blastulae of sea urchin, Hemicentrotus pulcherrimus, by means of 1 M KCl-extraction, phosphocellulose, DEAE-cellulose, Sepharose CL-6B, and double-stranded DNA cellulose column chromatography. The purified DNA ligase had a molecular weight of 80,000 (determined by Sephadex G-150) and a sedimentation coefficient of 4.1S (by glycerol gradient centrifugation). The purified enzyme required ATP and Mg2+ (or Mn2+) as cofactors for activity, and was inhibited by N-ethylmaleimide. Apparent Km values for ATP, Mg2+, and Mn2+ were 4 microM, 2.7 mM, and 0.3 mM, respectively.
D000818:Animals; D011088:DNA Ligases / Q000302:isolation & purification* / Q000378:metabolism; D004625:Embryo, Nonmammalian / Q000201:enzymology; D005033:Ethylmaleimide / Q000494:pharmacology; D005260:Female; D007700:Kinetics; D008274:Magnesium / Q000494:pharmacology; D008345:Manganese / Q000494:pharmacology; D008970:Molecular Weight; D011118:Polynucleotide Ligases / Q000302:isolation & purification*; D012617:Sea Urchins / Q000196:embryology*
D008345:Manganese; D011088:DNA Ligases; D011118:Polynucleotide Ligases; D008274:Magnesium; D005033:Ethylmaleimide