Tahirov Tahir H, Temiakov Dmitry, Anikin Michael, Patlan Vsevolod, McAllister William T, Vassylyev Dmitry G, Yokoyama Shigeyuki
High Throughput Factory, RIKEN Harima Institute at SPring-8, 1-1-1 Kouto, Mikazuki-cho, Sayo, Hyogo 679-5148, Japan.
Nature. 2002 Nov 7;420(6911):43-50. doi: 10.1038/nature01129. Epub 2002 Oct 9.
The single-subunit bacteriophage T7 RNA polymerase carries out the transcription cycle in an identical manner to that of bacterial and eukaryotic multisubunit enzymes. Here we report the crystal structure of a T7 RNA polymerase elongation complex, which shows that incorporation of an 8-base-pair RNA-DNA hybrid into the active site of the enzyme induces a marked rearrangement of the amino-terminal domain. This rearrangement involves alternative folding of about 130 residues and a marked reorientation (about 130 degrees rotation) of a stable core subdomain, resulting in a structure that provides elements required for stable transcription elongation. A wide opening on the enzyme surface that is probably an RNA exit pathway is formed, and the RNA-DNA hybrid is completely buried in a newly formed, deep protein cavity. Binding of 10 base pairs of downstream DNA is stabilized mostly by long-distance electrostatic interactions. The structure implies plausible mechanisms for the various phases of the transcription cycle, and reveals important structural similarities with the multisubunit RNA polymerases.
单亚基噬菌体T7 RNA聚合酶进行转录循环的方式与细菌和真核生物的多亚基酶相同。在此我们报道了T7 RNA聚合酶延伸复合物的晶体结构，该结构表明，一个8碱基对的RNA-DNA杂交体掺入酶的活性位点会诱导氨基末端结构域发生显著重排。这种重排涉及约130个残基的交替折叠以及一个稳定的核心亚结构域的显著重新定向（约130度旋转），从而形成一种为稳定转录延伸提供所需元件的结构。在酶表面形成了一个可能是RNA出口途径的宽开口，并且RNA-DNA杂交体完全埋藏在一个新形成的、较深的蛋白质腔中。下游10个碱基对DNA的结合主要通过长距离静电相互作用得以稳定。该结构暗示了转录循环各个阶段的合理机制，并揭示了与多亚基RNA聚合酶重要的结构相似性。
