Functional Interplay between the Jaw Domain of Bacterial RNA Polymerase and Allele-specific Residues in the Product RNA-binding Pocket

Ederth, Josefine Lundberg; Mooney, Rachel A.; Isaksson, Leif A.; Landick, Robert

doi:10.1016/j.jmb.2005.11.080

Cited by 33 publications

(29 citation statements)

References 64 publications

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“…Wild-type ECs and reduced F937–736 ECs synthesized the 78-nt RNA in 2–6 s at 37°C, 1 mM each NTP, suggesting an average transcription rate of 7–20 s −1 . This slower overall rate relative to the single- or double-turnover assays (Figure 2; 17) is comparable to prior observations for processive transcription under similar conditions (42,43) and reflects template positions at which translocation or catalysis may be slower than GTP addition on Scaffold GTP or at which pausing occurs. In reactions containing crosslinked F937–736 ECs, a distinct, slower elongating species is evident (red boxes, Supplementary Figure S1) with an average elongation rate of ∼1 s −1 .…”

Section: Resultssupporting

confidence: 87%

Trigger-helix folding pathway and SI3 mediate catalysis and hairpin-stabilized pausing by Escherichia coli RNA polymerase

Windgassen

Mooney

Nayak

et al. 2014

Nucleic Acids Research

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The conformational dynamics of the polymorphous trigger loop (TL) in RNA polymerase (RNAP) underlie multiple steps in the nucleotide addition cycle and diverse regulatory mechanisms. These mechanisms include nascent RNA hairpin-stabilized pausing, which inhibits TL folding into the trigger helices (TH) required for rapid nucleotide addition. The nascent RNA pause hairpin forms in the RNA exit channel and promotes opening of the RNAP clamp domain, which in turn stabilizes a partially folded, paused TL conformation that disfavors TH formation. We report that inhibiting TH unfolding with a disulfide crosslink slowed multiround nucleotide addition only modestly but eliminated hairpin-stabilized pausing. Conversely, a substitution that disrupts the TH folding pathway and uncouples establishment of key TH–NTP contacts from complete TH formation and clamp movement allowed rapid catalysis and eliminated hairpin-stabilized pausing. We also report that the active-site distal arm of the TH aids TL folding, but that a 188-aa insertion in the Escherichia coli TL (sequence insertion 3; SI3) disfavors TH formation and stimulates pausing. The effect of SI3 depends on the jaw domain, but not on downstream duplex DNA. Our results support the view that both SI3 and the pause hairpin modulate TL folding in a constrained pathway of intermediate states.

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Section: Resultssupporting

confidence: 87%

Trigger-helix folding pathway and SI3 mediate catalysis and hairpin-stabilized pausing by Escherichia coli RNA polymerase

Windgassen

Mooney

Nayak

et al. 2014

Nucleic Acids Research

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“…with a study suggesting a genetic link between residues in the β′ jaw and the active site of the RNAP (20).…”

Section: Discussionmentioning

confidence: 90%

T7 phage protein Gp2 inhibits the Escherichia coli RNA polymerase by antagonizing stable DNA strand separation near the transcription start site

Cámara¹,

Liu

Reynolds³

et al. 2010

Proc. Natl. Acad. Sci. U.S.A.

106

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Infection of Escherichia coli by the T7 phage leads to rapid and selective inhibition of the host RNA polymerase (RNAP)-a multisubunit enzyme responsible for gene transcription-by a small (∼7 kDa) phage-encoded protein called Gp2. Gp2 is also a potent inhibitor of E. coli RNAP in vitro. Here we describe the first atomic resolution structure of Gp2, which reveals a distinct run of surfaceexposed negatively charged amino acid residues on one side of the molecule. Our comprehensive mutagenesis data reveal that two conserved arginine residues located on the opposite side of Gp2 are important for binding to and inhibition of RNAP. Based on a structural model of the Gp2-RNAP complex, we propose that inhibition of transcription by Gp2 involves prevention of RNAPpromoter DNA interactions required for stable DNA strand separation and maintenance of the "transcription bubble" near the transcription start site, an obligatory step in the formation of a transcriptionally competent promoter complex.gene protein 2 | promoter melting | inhibitor

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“…A functional interplay between the β′ jaw and other mobile modules of the RNAP is not without precedent. A recent report by Ederth et al 18 suggests that the β′ jaw has an effect on the active centre of the RNAP through interactions with an active centre-proximal mobile element referred to as the β′ trigger loop. Thus, in light of our current results with the β′ clamp mutant Eσ 54 , it seems reasonable to propose that the contribution of the β′ jaw to open promoter complex stability involves an interplay with the β′ clamp.…”

Section: Discussionmentioning

confidence: 99%

Interplay between the β′ Clamp and the β′ Jaw Domains during DNA Opening by the Bacterial RNA Polymerase at σ54-dependent Promoters

Wigneshweraraj

Savalia

Severinov

et al. 2006

Journal of Molecular Biology

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Functional Interplay between the Jaw Domain of Bacterial RNA Polymerase and Allele-specific Residues in the Product RNA-binding Pocket

Cited by 33 publications

References 64 publications

Trigger-helix folding pathway and SI3 mediate catalysis and hairpin-stabilized pausing by Escherichia coli RNA polymerase

Trigger-helix folding pathway and SI3 mediate catalysis and hairpin-stabilized pausing by Escherichia coli RNA polymerase

T7 phage protein Gp2 inhibits the Escherichia coli RNA polymerase by antagonizing stable DNA strand separation near the transcription start site

Interplay between the β′ Clamp and the β′ Jaw Domains during DNA Opening by the Bacterial RNA Polymerase at σ54-dependent Promoters

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