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High-throughput Purification of Affinity-tagged Recombinant Proteins
Published on: August 26, 2012
用细菌RNA聚合酶进行转录延长的结构基础
Dmitry G Vassylyev1, Marina N Vassylyeva, Anna Perederina
1Department of Biochemistry and Molecular Genetics, University of Alabama at Birmingham, Schools of Medicine and Dentistry, 402B Kaul Genetics Building, 720 20th Street South, Birmingham, Alabama 35294, USA. dmitry@uab.edu
Nature
|June 22, 2007
概括
RNA聚合酶延长复合体 (EC) 结构揭示了它如何沿着DNA移动以合成RNA. 这为RNA聚合酶暂停和终止机制提供了洞察力.
科学领域:
- 分子生物学分子生物学
- 结构生物学 结构生物学
- 生物化学 生物化学
背景情况:
- RNA聚合酶延长复合体 (EC) 对于基因转录至关重要,可实现快速和过程性RNA合成.
- 了解欧洲共同体的结构动态是阐明转录机制和监管的关键.
研究的目的:
- 为了确定Thermus thermophilus EC.的高分辨率结构.
- 阐明RNA聚合酶过程性和调节的结构基础.
主要方法:
- 使用X射线晶体学以2.5A分辨率获得EC结构.
主要成果:
- 该结构捕获了一个转移后的中间体,其DNA模板可以在活性部位访问.
- 基因链分离发生在活性部位的下游,限制基质一次结合到一个.
- RNA/DNA杂交和移位RNA与特定的蛋白质元素相互作用,这表明了RNA移位机制.
- RNA退出通道构造提供了对依赖于发针的暂停和终止的见解.
结论:
- 高分辨率结构为EC在延长过程中提供了详细的分子视图.
- 结构特征解释了基质的可访问性,RNA的位移以及潜在的暂停/终止机制.
相关概念视频
Bacterial RNA Polymerase
Unlike eukaryotes, bacteria use a single RNA Polymerase (RNAP) to transcribe all genes. The different subunits of bacterial RNAPhave distinct functions. The multisubunit structure of the bacterial RNAP helps the enzyme to maintain catalytic function, facilitate assembly, interact with DNA and RNA, and self-regulate its activity.
In most genes, the transcription site is a single base present upstream of the coding sequence. Though RNAP is a catalytically efficient enzyme, it does not recognize...
In most genes, the transcription site is a single base present upstream of the coding sequence. Though RNAP is a catalytically efficient enzyme, it does not recognize...
Transcription Elongation Factors
Transcription elongation is a dynamic process that alters depending upon the sequence heterogeneity of the DNA being transcribed. Hence, it is not surprising that the elongation complex's composition also varies along the way while transcribing a gene.
The transcription elongation is regulated via pausing of RNA polymerase on several occasions during transcription. In bacteria, these halts are necessary because the transcription of DNA into mRNA is coupled to the translation of that mRNA into a...
The transcription elongation is regulated via pausing of RNA polymerase on several occasions during transcription. In bacteria, these halts are necessary because the transcription of DNA into mRNA is coupled to the translation of that mRNA into a...
Bacterial Transcription
RNA polymerase (RNAP) carries out DNA-dependent RNA synthesis in both bacteria and eukaryotes. Bacteria do not have a membrane-bound nucleus. So, transcription and translation occur simultaneously, on the same DNA template.
Transcription can be divided into three main stages, each involving distinct DNA sequences to guide the polymerase. These are:
Transcription can be divided into three main stages, each involving distinct DNA sequences to guide the polymerase. These are:
Transcription Initiation
Initiation is the first step of transcription in eukaryotes. Prokaryotic RNA Polymerase (RNAP) can bind to the template DNA and start transcribing. On the other hand, transcription in eukaryotes requires additional proteins, called transcription factors, to first bind to the promoter region in the DNA template. This binding helps recruit the specific RNAP that can assemble on the DNA and start transcription.
The promoters and enhancers and their accessory proteins allow tight regulation of...
The promoters and enhancers and their accessory proteins allow tight regulation of...
Bacterial RNA Polymerase
Unlike eukaryotes, bacteria use a single RNA Polymerase (RNAP) to transcribe all genes. The different subunits of bacterial RNAPhave distinct functions. The multisubunit structure of the bacterial RNAP helps the enzyme to maintain catalytic function, facilitate assembly, interact with DNA and RNA, and self-regulate its activity.
In most genes, the transcription site is a single base present upstream of the coding sequence. Though RNAP is a catalytically efficient enzyme, it does not recognize...
In most genes, the transcription site is a single base present upstream of the coding sequence. Though RNAP is a catalytically efficient enzyme, it does not recognize...
Transcription Elongation Factors
Transcription elongation is a dynamic process that alters depending upon the sequence heterogeneity of the DNA being transcribed. Hence, it is not surprising that the elongation complex's composition also varies along the way while transcribing a gene.
The transcription elongation is regulated via pausing of RNA polymerase on several occasions during transcription. In bacteria, these halts are necessary because the transcription of DNA into mRNA is coupled to the translation of that mRNA into a...
The transcription elongation is regulated via pausing of RNA polymerase on several occasions during transcription. In bacteria, these halts are necessary because the transcription of DNA into mRNA is coupled to the translation of that mRNA into a...

