相关实验视频
Updated: Jul 15, 2026

07:44
High-throughput Purification of Affinity-tagged Recombinant Proteins
Published on: August 26, 2012
转录延长因子GreB与细菌RNA聚合酶结合的结构和功能
Natacha Opalka1, Mark Chlenov, Pablo Chacon
1The Rockefeller University, 1230 York Avenue, New York, NY 10021, USA.
Cell
|August 14, 2003
概括
细菌的Gre因子 (GreA和GreB) 通过触发转录分离来增强RNA聚合酶转录. 结构分析揭示了希腊国家集团 (GreB) 的情况.
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:
- 细菌的Gre因子 (GreA和GreB) 是重要的转录延长因子.
- 它们刺激一种内在RNA聚合酶 (RNAP) 的内核分解活性,以切割新生的转录.
- 这一活动对于克服转录障碍和保持基因组稳定性至关重要.
研究的目的:
- 阐明GreB与RNAP活动相互作用和刺激的结构机制.
- 通过格雷因子介导的转录分离过程提供原子层次的洞察力.
主要方法:
- 低温电子显微镜 (低温电子显微镜) 分析了与GreB结合的*Escherichia coli*核心RNAP.
- 螺旋晶体的图像处理以达到15 Å的分辨率.
- 将RNAP和GreB的高分辨率晶体结构安装到冷EM图中.
- 针对位点的突变发生,以调查特定的GreB残留物的作用.
主要成果:
- 一个RNAP-GreB复合体的结构模型,揭示了GreB的N-终端线圈-线圈域延伸到RNAP活性位点通道.
- 确定在GreB卷轴的尖端保存的酸性残留物对于催化至关重要.
- 该模型解释了先前观察到的Gre因子活动,并为转录切割提供了机制基础.
结论:
- GreB通过其卷轴-卷轴域直接与RNAP活性部位相互作用,促进转录切割.
- 在GreB上保存的酸性残留物对于其在修改RNAP活性位点的催化功能至关重要.
- 本研究提供了对Gre因子介导的转录调节的详细结构理解.
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...

