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首页> 外文期刊>Journal of bacteriology >Molecular genetic analysis of a prokaryotic transcriptional coactivator: functional domains of the bacteriophage T4 gene 33 protein.
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Molecular genetic analysis of a prokaryotic transcriptional coactivator: functional domains of the bacteriophage T4 gene 33 protein.

机译：原核转录共激活子的分子遗传分析：噬菌体T4基因33蛋白的功能域。

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摘要

The bacteriophage T4 gene 33 encodes a small, acidic RNA polymerase-binding protein that mediates enhancement of transcriptional initiation at T4 late promoters by the T4 DNA replication accessory proteins. A set of nested deletions in the gene 33 open reading frame was constructed by oligonucleotide site-directed mutagenesis. The resulting variant gene 33 proteins were radiolabeled during overexpression employing a T7 RNA polymerase-based system and substantially purified. Each variant was analyzed for three properties of gp33: RNA polymerase binding activity, ability to mediate enhancer-dependent transcriptional activation, and repression of unenhanced transcription. Two separate regions of gp33 were required to form stable complexes with RNA polymerase, whereas the extreme carboxyl terminus of gp33 was essential for mediating late gene activation. Variant gene 33 proteins lacking the carboxyl terminus nevertheless repressed nonenhanced transcription, demonstrating that the functional domains required for transcriptional activation and repression of unenhanced transcription are separable. The possible roles of gp33 in mediating late gene expression are discussed in the light of the identification of these functional domains.
机译：噬菌体T4基因33编码小的酸性RNA聚合酶结合蛋白，其通过T4 DNA复制辅助蛋白介导在T4晚期启动子处转录起始的增强。通过寡核苷酸定点诱变构建基因33开放阅读框中的一组嵌套缺失。使用基于T7 RNA聚合酶的系统在过表达过程中对所得变异基因33蛋白进行放射性标记，并进行基本纯化。分析了每个变体的gp33的三个特性：RNA聚合酶结合活性，介导增强子依赖性转录激活的能力以及抑制未增强转录的能力。 gp33的两个独立区域需要与RNA聚合酶形成稳定的复合物，而gp33的极端羧基末端对于介导晚期基因激活至关重要。缺少羧基末端的变异基因33蛋白仍然抑制了未增强的转录，表明转录激活和抑制未增强的转录所需的功能域是可分离的。根据这些功能域的鉴定，讨论了gp33在介导晚期基因表达中的可能作用。

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