The RNA polymerase II trigger loop functions in substrate selection and is directly targeted by α-amanitin
The RNA polymerase II trigger loop functions in substrate selection and is directly targeted by α-amanitin
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DOI：
10.1016/j.molcel.2008.04.023
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发表时间：
2008-06-06
期刊：
影响因子：
16
通讯作者：
Kornberg, Roger D.
中科院分区：
文献类型：
--
作者：
Kaplan, Craig D.;Larsson, Karl-Magnus;Kornberg, Roger D.
结构、生化和遗传学研究表明，多亚基 RNA 聚合酶的移动元件触发环 (TL) 在催化中发挥着关键作用，并且可以成为抗生素抑制剂的靶点。在这里，我们提供了酿酒酵母 RNA 聚合酶 II (Pol II) TL 参与底物选择的证据。 Pol II TL 内的氨基酸取代优先改变底物使用和酶保真度，a-鹅膏蕈碱对转录的抑制也是如此。最后，TL 中 His1085 的取代特异性地使 Pol II 对 α-鹅膏蕈碱具有高度抗性，表明 His1085 和 α-鹅膏蕈碱之间存在功能性相互作用，这一相互作用得到了 α-鹅膏蕈碱-Pol II 晶体结构重新精炼的支持。我们认为，α-鹅膏蕈碱抑制的 Pol II 延伸缓慢且底物选择性降低，是由于 α-鹅膏蕈碱对 TL 的直接干扰造成的。
Structural, biochemical, and genetic studies have led to proposals that a mobile element of multisubunit RNA polymerases, the Trigger Loop (TL), plays a critical role in catalysis and can be targeted by antibiotic inhibitors. Here we present evidence that the Saccharomyces cerevisiae RNA Polymerase II (Pol II) TL participates in substrate selection. Amino acid substitutions within the Pol II TL preferentially alter substrate usage and enzyme fidelity, as does inhibition of transcription by a-amanitin. Finally, substitution of His1085 in the TL specifically renders Pol II highly resistant to alpha-amanitin, indicating a functional interaction between His1085 and a-amanitin that is supported by rerefinement of an alpha-amanitin-Pol II crystal structure. We propose that a-amanitin-inhibited Pol II elongation, which is slow and exhibits reduced substrate selectivity, results from direct oc-amanitin interference with the TL.