In vitro initial attachment of HIV-1 integrase to viral ends: control of the DNA specific interaction by the oligomerization state
In vitro initial attachment of HIV-1 integrase to viral ends: control of the DNA specific interaction by the oligomerization state
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DOI：
10.1093/nar/gkn796
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发表时间：
2008-12-01
影响因子：
14.9
通讯作者：
Parissi, V.
中科院分区：
文献类型：
--
作者：
Lesbats, P.;Metifiot, M.;Parissi, V.
HIV-1整合酶（IN）寡聚化和DNA识别是整合反应后续事件的关键步骤。最近的进展描述了在体外整合过程中包括二聚体和四聚体的稳定的中间复合物的参与，但最初的附着事件和病毒末端的IN定位尚不清楚。为了确定不同的IN寡聚复合物在这些早期步骤中的作用，我们进行了体外功能分析，比较具有不同寡聚化特性的IN制剂。我们证明，在体外IN协调整合活性的长DNA基板上含有特定的病毒和非特异性DNA序列是高度依赖于结合的预形成的二聚体的病毒末端。此外，我们表明，结合到非特异性DNA的IN单体也可以折叠成功能上不同的寡聚复合物，显示非特异性双链DNA断裂活性，而不是众所周知的单链切割催化相关的IN。我们的研究结果表明，有效形成的活性整合复合物高度需要早期正确定位的单体整合酶或直接结合的病毒末端的预制二聚体。总之，数据表明IN寡聚化控制酶特异性和活性。
HIV-1 integrase (IN) oligomerization and DNA recognition are crucial steps for the subsequent events of the integration reaction. Recent advances described the involvement of stable intermediary complexes including dimers and tetramers in the in vitro integration processes, but the initial attachment events and IN positioning on viral ends are not clearly understood. In order to determine the role of the different IN oligomeric complexes in these early steps, we performed in vitro functional analysis comparing IN preparations having different oligomerization properties. We demonstrate that in vitro IN concerted integration activity on a long DNA substrate containing both specific viral and nonspecific DNA sequences is highly dependent on binding of preformed dimers to viral ends. In addition, we show that IN monomers bound to nonspecific DNA can also fold into functionally different oligomeric complexes displaying nonspecific double-strand DNA break activity in contrast to the well known single strand cut catalyzed by associated IN. Our results imply that the efficient formation of the active integration complex highly requires the early correct positioning of monomeric integrase or the direct binding of preformed dimers on the viral ends. Taken together the data indicates that IN oligomerization controls both the enzyme specificity and activity.