相关实验视频
Updated: Jul 20, 2026

07:44
High-throughput Purification of Affinity-tagged Recombinant Proteins
Published on: August 26, 2012
概括
大肠杆菌RNA聚合酶通过17个基对解DNA,这是转录过程中的常数. 这表明酶活动,解酶和回酶,促进DNA螺旋环穿越和RNA转录位移.
科学领域:
- 分子生物学分子生物学
- 生物化学 生物化学
- 遗传学 是一个遗传学.
背景情况:
- DNA解是转录启动和延长的一个关键步骤.
- 通过RNA聚合酶操纵DNA螺旋的精确机制仍在调查中.
研究的目的:
- 为了量化埃舍里希亚大肠杆菌RNA聚合酶的DNA解.
- 阐明DNA解在转录启动和延长中的作用.
- 提出RNA聚合酶沿DNA运动的模型.
主要方法:
- 对二进制,初始化和三进制复合体的DNA解角的测量.
- 在猿人病毒40DNA上使用独特的促进子序列.
- 在37°C的温度下进行转录检测.
主要成果:
- 所有复合体都表现出一致的DNA解角为17±1个基对 (580°±30°).
- 在转录过程中,DNA解是酶稳定并维持的.
- 在延长过程中没有观察到解线角度的异质性.
结论:
- 建议在RNA聚合酶中存在"解酶"和"重酶"活动,用于DNA穿越.
- 建议一个模型,其中解酶解DNA,反酶取代RNA并重新封闭螺旋.
- 假设RNA-DNA杂交的形成是聚合酶暂停和终止的机制.
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 Attenuation in Prokaryotes
Transcriptional attenuation occurs when RNA transcription is prematurely terminated due to the formation of a terminator mRNA hairpin structure. Bacteria use these hairpins to regulate the transcription process and control the synthesis of several amino acids including histidine, lysine, threonine, and phenylalanine. Transcription attenuation takes place in the non-coding regions of mRNA.
There are several different mechanisms used to attenuate transcription. In ribosome mediated...
There are several different mechanisms used to attenuate transcription. In ribosome mediated...
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 in Prokaryotes
Transcription is a highly regulated process that converts genetic information into RNA molecules. The transcription cycle is divided into three key stages: initiation, elongation, and termination, each driven by specific molecular mechanisms.Initiation of TranscriptionIn bacteria, transcription begins when the RNA polymerase core enzyme associates with a sigma factor to form a holoenzyme. For example, the E. coli sigma factor called σ70 forms a holoenzyme, which recognizes the -10 (Pribnow box)...

