A DNA helicase activity is associated with an MCM4, -6, and -7 protein complex
A DNA helicase activity is associated with an MCM4, -6, and -7 protein complex
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DOI：
10.1074/jbc.272.39.24508
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发表时间：
1997-09-26
影响因子：
4.8
通讯作者：
Ishimi, Y
中科院分区：
文献类型：
--
作者：
Ishimi, Y
6种微小染色体维持(MCM)蛋白的中心结构域都有DNA依赖的ATPase基序，从酵母到哺乳动物都是保守的。本课题组利用组蛋白-琼脂糖柱层析(Ishimi，Y.，Ichinose，S.，Omori，A.，Sato K.和Kimura，H.(1996)J.Biol)从HeLa细胞中纯化了由MCM2，-4(CDC21)，-6(Mis5)和-7(CDC47)蛋白组成的MCM蛋白复合体。化学。271、24115-24122)。研究表明，ATPase活性和DNA解旋酶活性都与MCM蛋白复合体有关，ATPase活性和DNA解旋酶活性都与MCM蛋白复合体有关，ATPase和DNA解旋酶活性都与600 kDa的MCM4、-6和-7蛋白复合体共纯化，MCM蛋白复合体被抗MCM4抗体免疫耗尽，导致DNA解旋酶活性严重降低。DNA解旋酶对DNA片段的置换表明，DNA解旋酶沿单链DNA沿3‘-5’方向迁移，DNA解旋酶活性仅在水解性ATP或bATP存在时才能检测到。这些结果表明，该解旋酶可能作为DNA解旋酶参与了DNA复制的启动。
All six minichromosome maintenance (MCM) proteins have DNA-dependent ATPase motifs in the central domain which is conserved from yeast to mammals. Our group purified MCM protein complexes consisting of MCM2, -4 (Cdc21), -6 (Mis5), and -7 (CDC47) proteins from HeLa cells by using histone-Sepharose column chromatography (Ishimi, Y., Ichinose, S., Omori, A., Sato K., and Kimura, H. (1996) J. Biol. Chem. 271, 24115-24122). The present study revealed that both ATPase activity and DNA helicase activity that displaces oligonucleotides annealed to single-stranded circular DNA are associated with an MCM protein complex, Both ATPase and DNA helicase activities were co-purified with a 600-kDa protein complex that is consisted of equal amounts of MCM4, -6, and -7 proteins, An immunodepletion of the MCM protein complex from the purified fraction using anti-MCM4 antibody resulted in the severe reduction of the DNA helicase activity. Displacement of DNA fragments by the DNA helicase suggested that it migrated along single-stranded DNA in the 3' to 5' direction, and the DNA helicase activity was detected only in the presence of hydrolyzable ATP or bATP. These results suggest that this helicase may be involved in the initiation of DNA replication as a DNA unwinding enzyme.