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首页> 外文期刊>Molecular and Cellular Biology >Two distinct domains in the yeast transcription factor IID and evidence for a TATA box-induced conformational change.
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Two distinct domains in the yeast transcription factor IID and evidence for a TATA box-induced conformational change.

机译：酵母转录因子IID中的两个不同域和TATA盒诱导的构象变化的证据。

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Transcription factor IID from Saccharomyces cerevisiae (YIID) binds the TATA box element present in most RNA polymerase II promoters. In this work, partial proteolysis was used as a biochemical probe of YIID structure. YIID consists of a protease-sensitive amino terminus and a highly stable, protease-resistant carboxy-terminal core. The cleavage sites of the predominant chymotrypsin- and trypsin-derived fragments were mapped to amino acid residues 40 to 41 and 48 to 49, respectively, by amino-terminal peptide sequencing. Removal of the amino terminus resulted in a dramatic increase in the ability of YIID to form a stable complex with DNA during gel electrophoresis mobility shift assays and a two- to fourfold increase in DNA-binding affinity, as assayed by DNase I footprinting analysis. The carboxy-terminal 190-amino-acid core was competent for transcription in vitro and was similar in activity to native YIID. DNA containing a TATA element induced hypersensitive sites in the amino-terminal domain and stabilized the core domain to further proteolytic attack. Native YIID did not bind to a TATA box at 0 degrees C, whereas the carboxy-terminal DNA-binding domain did. These results suggest that YIID undergoes a conformational change upon binding to a TATA box. Southern blotting showed that the carboxy-terminal domain is highly conserved, while the amino-terminal domain diverged rapidly in evolution, even between closely related budding yeasts.
机译：来自酿酒酵母（YSID）的转录因子IID与大多数RNA聚合酶II启动子中存在的TATA盒元件结合。在这项工作中，部分蛋白水解被用作YIID结构的生化探针。 YIID由蛋白酶敏感的氨基末端和高度稳定，耐蛋白酶的羧基末端核心组成。胰凝乳蛋白酶和胰蛋白酶来源的主要片段的切割位点通过氨基末端肽测序分别定位到氨基酸残基40至41和48至49。如通过DNase I足迹分析所测定的，去除氨基末端导致YIID在凝胶电泳迁移率变动分析过程中与DNA形成稳定复合物的能力显着提高，并且DNA结合亲和力提高了2到4倍。羧基末端190个氨基酸的核心在体外具有转录能力，其活性与天然YIID相似。含有TATA元件的DNA会在氨基末端结构域中诱导超敏位点，并使核心结构域稳定，从而进一步发生蛋白水解攻击。天然YIID在0摄氏度时不与TATA盒结合，而羧基末端DNA结合域则与TATA盒结合。这些结果表明，当与TATA盒结合时，YIID发生构象变化。 Southern印迹表明，羧基末端结构域是高度保守的，而氨基末端结构域甚至在密切相关的出芽酵母之间在进化中迅速分歧。

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