scholarly journals Ureidothiophene inhibits interaction of bacterial RNA polymerase with –10 promotor element

2020 ◽  
Vol 48 (14) ◽  
pp. 7914-7923
Author(s):  
John Harbottle ◽  
Nikolay Zenkin
Keyword(s):  
Complex Formation ◽  
Rna Polymerase ◽  
Conformational Changes ◽  
Early Stage ◽  
Chemical Compounds ◽  
Initiation Factor ◽  
Promoter Element ◽  
Bacterial Transcription ◽  
Bacterial Rna ◽  
Novel Target

Abstract Bacterial RNA polymerase is a potent target for antibiotics, which utilize a plethora of different modes of action, some of which are still not fully understood. Ureidothiophene (Urd) was found in a screen of a library of chemical compounds for ability to inhibit bacterial transcription. The mechanism of Urd action is not known. Here, we show that Urd inhibits transcription at the early stage of closed complex formation by blocking interaction of RNA polymerase with the promoter –10 element, while not affecting interactions with –35 element or steps of transcription after promoter closed complex formation. We show that mutation in the region 1.2 of initiation factor σ decreases sensitivity to Urd. The results suggest that Urd may directly target σ region 1.2, which allosterically controls the recognition of –10 element by σ region 2. Alternatively, Urd may block conformational changes of the holoenzyme required for engagement with –10 promoter element, although by a mechanism distinct from that of antibiotic fidaxomycin (lipiarmycin). The results suggest a new mode of transcription inhibition involving the regulatory domain of σ subunit, and potentially pinpoint a novel target for development of new antibacterials.

scholarly journals The Core and Holoenzyme Forms of RNA Polymerase from Mycobacterium smegmatis

2018 ◽  
Vol 201 (4) ◽  
Author(s):  
Tomáš Kouba ◽  
Jiří Pospíšil ◽  
Jarmila Hnilicová ◽  
Hana Šanderová ◽  
Ivan Barvík ◽  
...  
Keyword(s):  
Electron Microscopy ◽  
Rna Polymerase ◽  
Conformational Changes ◽  
Mycobacterium Smegmatis ◽  
Drug Target ◽  
Conformational Dynamics ◽  
Three Dimensional ◽  
Cryo Electron Microscopy ◽  
Bacterial Rna ◽  
High Degree

ABSTRACT Bacterial RNA polymerase (RNAP) is essential for gene expression and as such is a valid drug target. Hence, it is imperative to know its structure and dynamics. Here, we present two as-yet-unreported forms of Mycobacterium smegmatis RNAP: core and holoenzyme containing σA but no other factors. Each form was detected by cryo-electron microscopy in two major conformations. Comparisons of these structures with known structures of other RNAPs reveal a high degree of conformational flexibility of the mycobacterial enzyme and confirm that region 1.1 of σA is directed into the primary channel of RNAP. Taken together, we describe the conformational changes of unrestrained mycobacterial RNAP. IMPORTANCE We describe here three-dimensional structures of core and holoenzyme forms of mycobacterial RNA polymerase (RNAP) solved by cryo-electron microscopy. These structures fill the thus-far-empty spots in the gallery of the pivotal forms of mycobacterial RNAP and illuminate the extent of conformational dynamics of this enzyme. The presented findings may facilitate future designs of antimycobacterial drugs targeting RNAP.

scholarly journals Engagement of Arginine Finger to ATP Triggers Large Conformational Changes in NtrC1 AAA+ ATPase for Remodeling Bacterial RNA Polymerase

Structure ◽  
2010 ◽  
Vol 18 (11) ◽  
pp. 1420-1430 ◽  
Author(s):  
Baoyu Chen ◽  
Tatyana A. Sysoeva ◽  
Saikat Chowdhury ◽  
Liang Guo ◽  
Sacha De Carlo ◽  
...  
Keyword(s):  
Rna Polymerase ◽  
Conformational Changes ◽  
Aaa Atpase ◽  
Bacterial Rna ◽  
Arginine Finger

scholarly journals Structural basis of RNA polymerase inhibition by viral and host factors

2021 ◽  
Vol 12 (1) ◽  
Author(s):  
Simona Pilotto ◽  
Thomas Fouqueau ◽  
Natalya Lukoyanova ◽  
Carol Sheppard ◽  
Soizick Lucas-Staat ◽  
...  
Keyword(s):  
Dna Binding ◽  
Rna Polymerase ◽  
Conformational Changes ◽  
Transcription Initiation ◽  
Environmental Changes ◽  
Initiation Factor ◽  
Structural Basis ◽  
Regulation Of Transcription ◽  
Domains Of Life ◽  
Sulfolobus Turreted Icosahedral Virus

AbstractRNA polymerase inhibition plays an important role in the regulation of transcription in response to environmental changes and in the virus-host relationship. Here we present the high-resolution structures of two such RNAP-inhibitor complexes that provide the structural bases underlying RNAP inhibition in archaea. The Acidianus two-tailed virus encodes the RIP factor that binds inside the DNA-binding channel of RNAP, inhibiting transcription by occlusion of binding sites for nucleic acid and the transcription initiation factor TFB. Infection with the Sulfolobus Turreted Icosahedral Virus induces the expression of the host factor TFS4, which binds in the RNAP funnel similarly to eukaryotic transcript cleavage factors. However, TFS4 allosterically induces a widening of the DNA-binding channel which disrupts trigger loop and bridge helix motifs. Importantly, the conformational changes induced by TFS4 are closely related to inactivated states of RNAP in other domains of life indicating a deep evolutionary conservation of allosteric RNAP inhibition.

scholarly journals The Phage-Encoded N-Acetyltransferase Rac Mediates Inactivation of Pseudomonas aeruginosa Transcription by Cleavage of the RNA Polymerase Alpha Subunit

Viruses ◽  
2020 ◽  
Vol 12 (9) ◽  
pp. 976 ◽  
Author(s):  
Pieter-Jan Ceyssens ◽  
Jeroen De Smet ◽  
Jeroen Wagemans ◽  
Natalia Akulenko ◽  
Evgeny Klimuk ◽  
...  
Keyword(s):  
Pseudomonas Aeruginosa ◽  
Rna Polymerase ◽  
Transcription Initiation ◽  
Alpha Subunit ◽  
Regulation Mechanism ◽  
Post Translational Modification ◽  
Α Subunit ◽  
Bacterial Transcription ◽  
Highly Active ◽  
Bacterial Rna

In this study, we describe the biological function of the phage-encoded protein RNA polymerase alpha subunit cleavage protein (Rac), a predicted Gcn5-related acetyltransferase encoded by phiKMV-like viruses. These phages encode a single-subunit RNA polymerase for transcription of their late (structure- and lysis-associated) genes, whereas the bacterial RNA polymerase is used at the earlier stages of infection. Rac mediates the inactivation of bacterial transcription by introducing a specific cleavage in the α subunit of the bacterial RNA polymerase. This cleavage occurs within the flexible linker sequence and disconnects the C-terminal domain, required for transcription initiation from most highly active cellular promoters. To achieve this, Rac likely taps into a novel post-translational modification (PTM) mechanism within the host Pseudomonas aeruginosa. From an evolutionary perspective, this novel phage-encoded regulation mechanism confirms the importance of PTMs in the prokaryotic metabolism and represents a new way by which phages can hijack the bacterial host metabolism.

scholarly journals RNA extension drives a stepwise displacement of an initiation-factor structural module in initial transcription

2020 ◽  
Vol 117 (11) ◽  
pp. 5801-5809 ◽  
Author(s):  
Lingting Li ◽  
Vadim Molodtsov ◽  
Wei Lin ◽  
Richard H. Ebright ◽  
Yu Zhang
Keyword(s):  
Rna Polymerase ◽  
Crystal Structures ◽  
Active Center ◽  
Transcription Initiation ◽  
Initiation Factor ◽  
Template Strand ◽  
Bacterial Transcription ◽  
Transcription Initiation Factor ◽  
Σ Factor ◽  
Nascent Rna

All organisms—bacteria, archaea, and eukaryotes—have a transcription initiation factor that contains a structural module that binds within the RNA polymerase (RNAP) active-center cleft and interacts with template-strand single-stranded DNA (ssDNA) in the immediate vicinity of the RNAP active center. This transcription initiation-factor structural module preorganizes template-strand ssDNA to engage the RNAP active center, thereby facilitating binding of initiating nucleotides and enabling transcription initiation from initiating mononucleotides. However, this transcription initiation-factor structural module occupies the path of nascent RNA and thus presumably must be displaced before or during initial transcription. Here, we report four sets of crystal structures of bacterial initially transcribing complexes that demonstrate and define details of stepwise, RNA-extension-driven displacement of the “σ-finger” of the bacterial transcription initiation factor σ. The structures reveal that—for both the primary σ-factor and extracytoplasmic (ECF) σ-factors, and for both 5′-triphosphate RNA and 5′-hydroxy RNA—the “σ-finger” is displaced in stepwise fashion, progressively folding back upon itself, driven by collision with the RNA 5′-end, upon extension of nascent RNA from ∼5 nt to ∼10 nt.

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