Analysis of M phase-specific phosphorylation of DNA topoisomerase II
Analysis of M phase-specific phosphorylation of DNA topoisomerase II
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DOI：
10.1074/jbc.271.35.21439
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发表时间：
1996-08-30
影响因子：
4.8
通讯作者：
Kikuchi, A
中科院分区：
文献类型：
--
作者：
Kimura, K;Nozaki, N;Kikuchi, A
在哺乳动物细胞中，DNA拓扑异构酶II（topo II）的两种同种型，topo II α和topo II β被磷酸化。topo II β的磷酸化在有丝分裂细胞中将其表观分子量从180 kDa改变至190 kDa，而topo II α仅轻微影响它（Kimura，K.，Nozaki，N.，西条，M.，菊池，A.，Ui，M，和Enomoto，T.（1994）J，Biol，Chem，269，24523-24526）。在此，我们检查了当细胞从M期进展到G（1）期时，每种topo II同种型的蛋白质和磷酸盐部分的稳定性。虽然其蛋白质部分保持完整，但在从有丝分裂阻滞释放后4小时内，75%的附着于topo II β的磷酸盐被除去。另一方面，35%的topo Ⅱ α蛋白和52%的附着磷酸盐消失。我们证实了在广泛的磷酸酶消化后，M期特异性磷酸化对两种topo Ⅱ亚型的催化活性没有特别的影响。我们还检查了两种亚型与细胞核或染色体的结合。从有丝分裂染色体，拓扑II β提取在低得多的浓度的NaCl比拓扑II α。
In mammalian cells, two isoforms of DNA topoisomerase II (topo II), topo II alpha and topo II beta, are phosphorylated, The phosphorylation of topo II beta changes its apparent molecular mass determined by SDS-polyacrylamide gel electrophoresis from 180 to 190 kDa in mitotic cells, whereas topo II alpha affects it only slightly (Kimura, K,, Nozaki, N,, Saijo, M,, Kikuchi, A., Ui, M,, and Enomoto, T. (1994) J, Biol, Chem, 269, 24523-24526), Here we examined the stability of the protein and the phosphate moiety of each topo II isoform, as the cells progressed from M to G(1) phase, While its protein moiety remained intact, 75% of the phosphates attached to topo II beta were removed within 4 h after release from mitotic block, On the other hand, 35% of topo II alpha protein and 52% of the attached phosphates disappeared, We verified that M phase-specific phosphorylation had no particular effect on the catalytic activities of both topo II isoforms after extensive phosphatase digestion, We also examined the binding of two isoforms to the nucleus or chromosomes, In logarithmically growing cells, both isoforms were extracted from nuclei at the same concentrations of NaCl, From the mitotic chromosomes, topo II beta was extracted at much lower concentrations of NaCl than topo II alpha.