Initiation of adenovirus DNA replication. I. Mechanism of action of a host protein required for replication of adenovirus DNA templates devoid of the terminal protein.
Initiation of adenovirus DNA replication. I. Mechanism of action of a host protein required for replication of adenovirus DNA templates devoid of the terminal protein.
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发表时间：
1988-07
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影响因子：
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通讯作者：
M. Kenny;L. A. Balogh;J. Hurwitz
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作者：
M. Kenny;L. A. Balogh;J. Hurwitz
与55-kDa末端蛋白共价连接的腺病毒（Ad）DNA的体外复制需要至少五种蛋白，包括80-kDa前末端蛋白、Ad DNA聚合酶、Ad DNA结合蛋白、核因子I和拓扑异构酶I。缺乏末端蛋白的Ad DNA模板的复制需要另外的蛋白，称为因子pL，其已经从未感染的HeLa细胞核中纯化（Guggenheimer，R.一、Nagata，K.，肯尼，M.，和Hurwitz，J.（1984）J.Biol.Chem.259，7815-7825）。已发现因子pL含有固有的5 ′-3 ′核酸外切酶活性。当缺乏末端蛋白的Ad DNA模板用因子pL预处理时，复制反应中对因子pL的需要被取消。构建含有Ad起始序列的合成部分双链体寡核苷酸模板，以确定在因子pL不存在下有活性的DNA分子的结构。这些实验表明，因子pL降解Ad起点的非模板（置换）链的5 '端，从而在模板链的3'端产生单链区域。这样的DNA在没有因子pL的情况下能够启动Ad DNA复制，但仍然依赖于核因子1。
The in vitro replication of adenovirus (Ad) DNA covalently attached to the 55-kDa terminal protein requires at least five proteins including the 80-kDa preterminal protein, the Ad DNA polymerase, the Ad DNA binding protein, nuclear factor I, and topoisomerase I. The replication of Ad DNA templates devoid of the terminal protein requires an additional protein, designated factor pL, which has been purified from uninfected HeLa cell nuclei (Guggenheimer, R. A., Nagata, K., Kenny, M., and Hurwitz, J. (1984) J. Biol. Chem. 259, 7815-7825). Factor pL has been found to contain an intrinsic 5'----3' exonuclease activity. When Ad DNA templates lacking the terminal protein were pretreated with factor pL, the requirement for factor pL in the replication reaction was abolished. Synthetic partially duplex oligonucleotide templates containing Ad origin sequences were constructed in order to determine the structure of the DNA molecules that are active in the absence of factor pL. These experiments indicated that factor pL degrades the 5'-end of the nontemplate (displaced) strand of the Ad origin thereby creating a single-stranded region at the 3'-end of the template strand. Such DNAs are competent for initiation of Ad DNA replication in the absence of factor pL but remain dependent on nuclear factor 1.