Im D S, Muzyczka N
Department of Microbiology, SUNY Stony Brook Medical School 11794.
Cell. 1990 May 4;61(3):447-57. doi: 10.1016/0092-8674(90)90526-k.
Genetic studies of adeno-associated virus (AAV) indicate that two AAV genes are required for viral DNA replication: the palindromic terminal repeat, which is the origin for DNA replication, and the rep gene, which codes for a family of at least four viral nonstructural proteins. To determine the biochemical function of the Rep proteins, we have purified the AAV Rep68 protein to apparent homogeneity. We find that it contains a site-specific and strand-specific endonuclease activity that specifically cuts the AAV origin at the terminal resolution site (TRS). The TRS endonuclease requires the presence of ATP for activity and becomes covalently attached to the 5' end at the cut site. In addition to the specific endonuclease activity, Rep68 also contains a DNA helicase activity. These results demonstrate that the large AAV Rep proteins have a direct role in AAV DNA replication; namely, they provide the activities required for the resolution of covalently joined AAV termini.
腺相关病毒（AAV）的遗传学研究表明，病毒DNA复制需要两个AAV基因：回文末端重复序列，它是DNA复制的起点；以及rep基因，它编码一个至少由四种病毒非结构蛋白组成的家族。为了确定Rep蛋白的生化功能，我们已将AAV Rep68蛋白纯化至表观均一性。我们发现它具有位点特异性和链特异性的内切核酸酶活性，可在末端分辨率位点（TRS）特异性切割AAV起源。TRS内切核酸酶的活性需要ATP的存在，并在切割位点与5'末端共价连接。除了特异性内切核酸酶活性外，Rep68还具有DNA解旋酶活性。这些结果表明，大型AAV Rep蛋白在AAV DNA复制中具有直接作用；即，它们提供了切割共价连接的AAV末端所需的活性。
