Highly specific aptamer trap for extremophilic RNA polymerases

IF 3.3 3区 生物学 Q2 BIOCHEMISTRY & MOLECULAR BIOLOGY
Ivan Petushkov , Andrey Feklistov , Andrey Kulbachinskiy
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引用次数: 0

Abstract

During transcription initiation, the holoenzyme of bacterial RNA polymerase (RNAP) specifically recognizes promoters using a dedicated σ factor. During transcription elongation, the core enzyme of RNAP interacts with nucleic acids mainly nonspecifically, by stably locking the DNA template and RNA transcript inside the main cleft. Here, we present a synthetic DNA aptamer that is specifically recognized by both core and holoenzyme RNAPs from extremophilic bacteria of the Deinococcus-Thermus phylum. The aptamer binds RNAP with subnanomolar affinities, forming extremely stable complexes even at high ionic strength conditions, blocks RNAP interactions with the DNA template and inhibits RNAP activity during transcription elongation. We propose that the aptamer binds at a conserved site within the downstream DNA-binding cleft of RNAP and traps it in an inactive conformation. The aptamer can potentially be used for structural studies to reveal RNAP conformational states, affinity binding of RNAP and associated factors, and screening of transcriptional inhibitors.

针对嗜极 RNA 聚合酶的高特异性适配体陷阱。
在转录启动过程中,细菌 RNA 聚合酶(RNAP)的全酶利用专用的 σ 因子特异性地识别启动子。在转录延伸过程中,RNAP 的核心酶主要通过将 DNA 模板和 RNA 转录本稳定地锁定在主裂隙内与核酸进行非特异性相互作用。在这里,我们展示了一种合成的DNA适配体,这种适配体能被来自嗜极细菌门(Deinococcus-Thermus)的核心和全酶RNAP特异性识别。这种适配体能以亚纳摩尔级的亲和力与 RNAP 结合,即使在高离子强度条件下也能形成极其稳定的复合物,阻断 RNAP 与 DNA 模板的相互作用,并在转录延伸过程中抑制 RNAP 的活性。我们认为,该配合物与 RNAP 下游 DNA 结合裂隙中的一个保守位点结合,并使其处于非活性构象。这种适配体可用于结构研究以揭示 RNAP 的构象状态、RNAP 与相关因子的亲和结合以及转录抑制剂的筛选。
本文章由计算机程序翻译,如有差异,请以英文原文为准。
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来源期刊
Biochimie
Biochimie 生物-生化与分子生物学
CiteScore
7.20
自引率
2.60%
发文量
219
审稿时长
40 days
期刊介绍: Biochimie publishes original research articles, short communications, review articles, graphical reviews, mini-reviews, and hypotheses in the broad areas of biology, including biochemistry, enzymology, molecular and cell biology, metabolic regulation, genetics, immunology, microbiology, structural biology, genomics, proteomics, and molecular mechanisms of disease. Biochimie publishes exclusively in English. Articles are subject to peer review, and must satisfy the requirements of originality, high scientific integrity and general interest to a broad range of readers. Submissions that are judged to be of sound scientific and technical quality but do not fully satisfy the requirements for publication in Biochimie may benefit from a transfer service to a more suitable journal within the same subject area.
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