Chen G, Stenlund A
Cold Spring Harbor Laboratory, New York 11724, USA.
J Virol. 1998 Apr;72(4):2567-76. doi: 10.1128/JVI.72.4.2567-2576.1998.
The bovine papillomavirus replication initiator protein E1 is an origin of replication (ori)-binding protein absolutely required for viral DNA replication. In the presence of the viral transcription factor E2, E1 binds to the ori and initiates DNA replication. To understand how the E1 initiator recognizes the ori and how E2 assists in this process, we have expressed and purified a 166-amino-acid fragment which corresponds to the minimal E1 DNA-binding domain (DBD). DNA binding studies using this protein demonstrate that the E1 DBD can bind to the palindromic E1 binding site in several forms but that binding of two monomers, each recognizing one half-site of the E1 palindrome, is the predominant form. This is reminiscent of the binding of the T-antigen DBD to the SV40 ori, and interestingly, the arrangement of E1 binding sites shows striking similarities to the arrangement of T-antigen binding sites in the SV40 ori even though the recognition sequences are unrelated. The E1 DBD is capable of interacting cooperatively with E2; however, the E2 DBD and not the E2 activation domain mediates this interaction. Furthermore, the E2 DBD stimulates binding of two monomers of the E1 DBD to the ori by binding cooperatively with one E1 monomer. Finally, we show that our results concerning the DNA-binding properties of the E1 DBD can be extended to full-length E1.
牛乳头瘤病毒复制起始蛋白E1是病毒DNA复制绝对必需的一种复制起点（ori）结合蛋白。在病毒转录因子E2存在的情况下，E1与ori结合并启动DNA复制。为了了解E1起始蛋白如何识别ori以及E2如何协助这一过程，我们表达并纯化了一个166个氨基酸的片段，该片段对应于最小的E1 DNA结合结构域（DBD）。使用该蛋白进行的DNA结合研究表明，E1 DBD可以以多种形式与回文E1结合位点结合，但两个单体的结合是主要形式，每个单体识别E1回文的一个半位点。这让人联想到T抗原DBD与SV40 ori的结合，有趣的是，尽管识别序列无关，但E1结合位点的排列与SV40 ori中T抗原结合位点的排列有显著相似性。E1 DBD能够与E2协同相互作用；然而，介导这种相互作用的是E2 DBD而不是E2激活结构域。此外，E2 DBD通过与一个E1单体协同结合来刺激E1 DBD的两个单体与ori的结合。最后，我们表明，我们关于E1 DBD DNA结合特性的结果可以扩展到全长E1。
