Lavigne M, Buc H
Unité de Physicochimie des Macromolécules Biologiques (CNRS URA 1773), Institut Pasteur, 75724 Paris Cedex 15, France.
J Mol Biol. 1999 Jan 22;285(3):977-95. doi: 10.1006/jmbi.1998.2367.
HIV-1 reverse transcriptase (RT) generally terminates plus strand DNA synthesis at the centre of the viral genome. The central termination sequence (CTS) contains two termination sites which are located at the 3' end of AnTm motifs. These motifs generate a global curvature of the DNA helix which correlates with termination of DNA synthesis. Here, we have characterized HIV-1 RT termination sites on different DNA sequences. Again, they are located at the 3' end of A-tracts. Using hydroxyl radicals as a probe of the width of the DNA helix, we have shown that RT termination sites are always located a few base-pairs downstream of a compressed minor groove. Mutations which relieve these compressions also abolish the termination events. The replacement of the adenine tracts by 2,6-diaminopurine tracts has a similar effect. Moreover, no termination site is observed on DNA sequences containing phased GC-tracts which curve the DNA helix but compress the major groove. The compression of the DNA minor groove and not necessarily the curved trajectory of the DNA is, therefore, responsible for termination of DNA synthesis at the CTS by HIV-1 RT. This conclusion is consistent with interpretation of other biochemical data on the processivity of HIV-1 RT, based on the structure of a DNA-enzyme complex.
HIV-1逆转录酶（RT）通常在病毒基因组的中心终止正链DNA合成。中央终止序列（CTS）包含两个终止位点，它们位于AnTm基序的3'端。这些基序会产生DNA螺旋的整体曲率，这与DNA合成的终止相关。在此，我们对不同DNA序列上的HIV-1 RT终止位点进行了表征。同样，它们位于A序列的3'端。使用羟基自由基作为DNA螺旋宽度的探针，我们发现RT终止位点总是位于压缩小沟下游的几个碱基对处。缓解这些压缩的突变也会消除终止事件。用2,6-二氨基嘌呤序列取代腺嘌呤序列具有类似的效果。此外，在含有使DNA螺旋弯曲但压缩大沟的相间GC序列的DNA序列上未观察到终止位点。因此，DNA小沟的压缩而非DNA的弯曲轨迹是HIV-1 RT在CTS处终止DNA合成的原因。这一结论与基于DNA-酶复合物结构对HIV-1 RT持续性的其他生化数据的解释一致。
