Crystal structure of bacterial RNA polymerase bound with a transcription inhibitor protein
Crystal structure of bacterial RNA polymerase bound with a transcription inhibitor protein
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DOI：
10.1038/nature09573
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发表时间：
2010-12-16
期刊：
影响因子：
64.8
通讯作者：
Yokoyama, Shigeyuki
中科院分区：
文献类型：
--
作者：
Tagami, Shunsuke;Sekine, Shun-ichi;Yokoyama, Shigeyuki
多亚基DNA依赖的RNA聚合酶(RNAP)是基因表达的主要转录酶。转录受多种转录因子的调控。GRE因子同源物1(Gfh1)，发现于Thermus属，是保守的细菌转录因子greA的紧密同源物，通过与RNAP(1-8)直接结合而抑制转录起始和延伸。Gfh1抑制转录的结构基础仍然不清楚，尽管RNAP和Gfh1的晶体结构已经分别确定(6-9)。本文报道了与Gfh1络合的嗜热嗜热菌RNAP的晶体结构。Gfh1的氨基末端螺旋结构域完全封闭了RNAP的两个中央模块之间形成的通道；该通道通常用于三磷酸核苷酸(NTP)进入催化部位。此外，螺旋结构域的尖端占据了NTPβ-γ磷酸结合部位。NTP进入通道被扩展，因为与以前报道的伸长复合体相比，中央模块相对于彼此‘棘轮’移动了类似7度。这种“棘轮状态”是另一种结构状态，由中央模块之间新获得的接触所定义。因此，Gfh1的形状适合维持RNAP处于棘轮状态。同时，棘轮作用扩大了核酸结合通道，并扭曲了连接中央模块的桥式螺旋。综上所述，目前的结果表明，Gfh1通过阻止NTP结合和冻结处于交替结构状态的RNAP来抑制转录。棘轮状态也可能与转录的其他方面有关，如RNAP易位和转录终止。
The multi-subunit DNA-dependent RNA polymerase (RNAP) is the principal enzyme of transcription for gene expression. Transcription is regulated by various transcription factors. Gre factor homologue 1 (Gfh1), found in the Thermus genus, is a close homologue of the well-conserved bacterial transcription factor GreA, and inhibits transcription initiation and elongation by binding directly to RNAP(1-8). The structural basis of transcription inhibition by Gfh1 has remained elusive, although the crystal structures of RNAP and Gfh1 have been determined separately(6-9). Here we report the crystal structure of Thermus thermophilus RNAP complexed with Gfh1. The amino-terminal coiled-coil domain of Gfh1 fully occludes the channel formed between the two central modules of RNAP; this channel would normally be used for nucleotide triphosphate (NTP) entry into the catalytic site. Furthermore, the tip of the coiled-coil domain occupies the NTP beta-gamma phosphate-binding site. The NTP-entry channel is expanded, because the central modules are 'ratcheted' relative to each other by similar to 7 degrees, as compared with the previously reported elongation complexes. This 'ratcheted state' is an alternative structural state, defined by a newly acquired contact between the central modules. Therefore, the shape of Gfh1 is appropriate to maintain RNAP in the ratcheted state. Simultaneously, the ratcheting expands the nucleic-acid-binding channel, and kinks the bridge helix, which connects the central modules. Taken together, the present results reveal that Gfh1 inhibits transcription by preventing NTP binding and freezing RNAP in the alternative structural state. The ratcheted state might also be associated with other aspects of transcription, such as RNAP translocation and transcription termination.