An Insertion in the Catalytic Trigger Loop Gates the Secondary Channel of RNA Polymerase

JOURNAL OF MOLECULAR BIOLOGY

卷:425

Article

Furman, Ran^1,2  Tsodikov, Oleg V.³  Wolf, Yuri I.⁴  Artsimovitch, Irina^1,2 

[1] Ohio State Univ, Dept Microbiol, Columbus, OH 43210 USA

[2] Ohio State Univ, Ctr RNA Biol, Columbus, OH 43210 USA

[3] Univ Michigan, Coll Pharm, Depatment Med Chem, Ann Arbor, MI 48105 USA

[4] NIH, Natl Ctr Biotechnol Informat, Natl Lib Med, Bethesda, MD 20892 USA

关键词: DksA;    Gre;    RNA polymerase;    secondary channel;    trigger loop;   

DOI  :  10.1016/j.jmb.2012.11.008

来源: Elsevier

【 摘 要 】

Escherichia coli DksA and GreB bind to RNA polymerase (RNAP), reaching inside the secondary channel, with similar affinities but have different cellular functions. DksA destabilizes promoter complexes whereas GreB facilitates RNA cleavage in arrested elongation complexes (ECs). Although the less abundant GreB may not interfere with DksA regulation during initiation, reports that DksA acts during elongation and termination suggest that it may exclude GreB from arrested complexes, potentially triggering genome instability. Here, we show that GreB does not compete with DksA during termination whereas DksA, even when present in several hundredfold molar excess, does not inhibit GreB-mediated cleavage of the nascent RNA. Our findings that DksA does not bind to backtracked or active ECs provide an explanation for the lack of DksA activity on most ECs that we reported previously, raising a question of what makes a transcription complex susceptible to DksA. Structural modeling suggests that i6, an insertion in the catalytic trigger loop, hinders DksA access into the channel, restricting DksA action to a subset of transcription complexes. In support of this hypothesis, we demonstrate that deletion of i6 permits DksA binding to ECs and that the distribution of DksA and i6 in bacterial genomes is strongly concordant. We hypothesize that DksA binds to transcription complexes in which i6 becomes mobile, for example, as a consequence of weakened RNAP interactions with the downstream duplex DNA. (C) 2012 Elsevier Ltd. All rights reserved.

【 授权许可】

Free