Takahashi M, Senshu M
FEBS Lett. 1987 Mar 23;213(2):345-52. doi: 10.1016/0014-5793(87)81520-x.
Embryos of Drosophila melanogaster contain two distinct DNA ligases (DNA ligase I and II). DNA ligase I was eluted at 0.2 M KCl and DNA ligase II at 0.6 M KCl on phosphocellulose column chromatography. The former was rich in early developing embryos and its activity decreased during embryonic development. The latter was found constantly throughout the developing stages of embryos. DNA ligase I existed in a cytoplasmic fraction and DNA ligase II is concentrated in nuclei. Both enzymes ligate 5'-phosphoryl and 3'-hydroxyl groups in oligo(dT) in the presence of poly(dA). DNA ligase II is also able to join oligo(dT)(poly(rA). Both enzymes require ATP and Mg2+ for activity. The Km for ATP is 2.7 X 10(-6) M for DNA ligase I, and 3.0 X 10(-5) M for DNA ligase II. DNA ligase I requires dithiothreitol and polyvinyl alcohol, but DNA ligase II does not. Both enzymes are inhibited in the presence of N-ethylmaleimide. DNA ligase I is active at a low salt concentration (0-30 mM KCl), but DNA ligase II is active at high salt concentrations (50-100 mM). DNA ligase I is more labile than DNA ligase II. The molecular masses of DNA ligase-AMP adducts were determined as 86 and 75 kDa for DNA ligase I, and as 70 (major protein) and 90 kDa (minor protein) for DNA ligase II under denaturing conditions. A sedimentation coefficient of 4.2 S was observed for DNA ligase II. Consequently, Drosophila DNA ligase I and II are quite similar to mammalian DNA ligase I and II. Drosophila DNA ligase I and a DNA ligase by B.A. Rabin et al. [(1986) J. Biol. Chem. 261, 10637-10645] seem to be the same enzyme.
黑腹果蝇胚胎含有两种不同的DNA连接酶（DNA连接酶I和II）。在磷酸纤维素柱层析中，DNA连接酶I在0.2M KCl浓度下被洗脱，而DNA连接酶II在0.6M KCl浓度下被洗脱。前者在早期发育胚胎中含量丰富，其活性在胚胎发育过程中降低。后者在胚胎发育的各个阶段都持续存在。DNA连接酶I存在于细胞质部分，而DNA连接酶II集中在细胞核中。在聚（dA）存在的情况下，两种酶都能连接寡聚（dT）中的5'-磷酸基和3'-羟基。DNA连接酶II也能够连接寡聚（dT）（聚（rA）。两种酶的活性都需要ATP和Mg2+。DNA连接酶I对ATP的Km值为2.7×10（-6）M，DNA连接酶II为3.0×10（-5）M。DNA连接酶I需要二硫苏糖醇和聚乙烯醇，而DNA连接酶II则不需要。两种酶在N-乙基马来酰亚胺存在时均被抑制。DNA连接酶I在低盐浓度（0-30mM KCl）下有活性，而DNA连接酶II在高盐浓度（50-100mM）下有活性。DNA连接酶I比DNA连接酶II更不稳定。在变性条件下，DNA连接酶I的DNA连接酶-AMP加合物的分子量测定为86和75kDa，DNA连接酶II的为70（主要蛋白）和90kDa（次要蛋白）。观察到DNA连接酶II的沉降系数为4.2S。因此，果蝇DNA连接酶I和II与哺乳动物DNA连接酶I和II非常相似。果蝇DNA连接酶I与B.A. Rabin等人[(1986) J. Biol. Chem. 261, 10637-10645]报道的一种DNA连接酶似乎是同一种酶。
