Folding and ligand recognition of the TPP riboswitch aptamer at single-molecule resolution

Haller, Andrea; Altman, Roger B.; Soulière, Marie F.; Blanchard, Scott C.; Micura, Ronald

doi:10.1073/pnas.1218062110

Cited by 115 publications

(170 citation statements)

References 49 publications

(89 reference statements)

Supporting

Mentioning

156

Contrasting

Order By: Relevance

“…For instance, in most smFRET studies, only a single structural perspective on riboswitch dynamics has been obtained (e.g., from just one FRET pair). The importance of applying multiple structural perspectives in smFRET imaging experiments is exemplified by the present investigation, prior studies of the TPP riboswitch aptamer (49), as well as more complex systems such as DNA helicase (50) and the bacterial ribosome (51,52). Such studies have the potential to uncover unexpected complexities in the structure and dynamics that would be difficult or impossible to discern through bulk techniques.…”

Section: Large Structurally Complex Riboswitches With Pseudoknots Andmentioning

confidence: 81%

Tuning a riboswitch response through structural extension of a pseudoknot

Soulière

Altman

Schwarz

et al. 2013

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

Self Cite

102

View full text Add to dashboard Cite

Structural and dynamic features of RNA folding landscapes represent critical aspects of RNA function in the cell and are particularly central to riboswitch-mediated control of gene expression. Here, using single-molecule fluorescence energy transfer imaging, we explore the folding dynamics of the preQ 1 class II riboswitch, an upstream mRNA element that regulates downstream encoded modification enzymes of queuosine biosynthesis. For reasons that are not presently understood, the classical pseudoknot fold of this system harbors an extra stem-loop structure within its 3′-terminal region immediately upstream of the Shine-Dalgarno sequence that contributes to formation of the ligand-bound state. By imaging ligand-dependent preQ 1 riboswitch folding from multiple structural perspectives, we reveal that the extra stem-loop strongly influences pseudoknot dynamics in a manner that decreases its propensity to spontaneously fold and increases its responsiveness to ligand binding. We conclude that the extra stem-loop sensitizes this RNA to broaden the dynamic range of the ON/OFF regulatory switch.SHAPE | single-molecule FRET | site-specific labeling | metabolite sensing RNAs | ligand recognition A variety of small metabolites have been found to regulate gene expression in bacteria, fungi, and plants via direct interactions with distinct mRNA folds (1-4). In this form of regulation, the target mRNA typically undergoes a structural change in response to metabolite binding (5-9). These mRNA elements have thus been termed "riboswitches" and generally include both a metabolite-sensitive aptamer subdomain and an expression platform. For riboswitches that regulate the process of translation, the expression platform minimally consists of a ribosomal recognition site [Shine-Dalgarno (SD)]. In the simplest form, the SD sequence overlaps with the metabolite-sensitive aptamer domain at its downstream end. Representative examples include the S-adenosylmethionine class II (SAM-II) (10) and the S-adenosylhomocysteine (SAH) riboswitches (11, 12), as well as prequeuosine class I (preQ 1 -I) and II (preQ 1 -II) riboswitches (13,14). The secondary structures of these four short RNA families contain a pseudoknot fold that is central to their gene regulation capacity. Although the SAM-II and preQ 1 -I riboswitches fold into classical pseudoknots (15, 16), the conformations of the SAH (17) and preQ 1 -II counterparts are more complex and include a structural extension that contributes to the pseudoknot architecture (14). Importantly, the impact and evolutionary significance of these "extra" stem-loop elements on the function of the SAH and preQ 1 -II riboswitches remain unclear.PreQ 1 riboswitches interact with the bacterial metabolite 7-aminomethyl-7-deazaguanine (preQ 1 ), a precursor molecule in the biosynthetic pathway of queuosine, a modified base encountered at the wobble position of some transfer RNAs (14). The general biological significance of studying the preQ 1 -II system stems from the fact that this gene-regulatory element is found...

show abstract

Section: Large Structurally Complex Riboswitches With Pseudoknots Andmentioning

confidence: 81%

Tuning a riboswitch response through structural extension of a pseudoknot

Soulière

Altman

Schwarz

et al. 2013

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

Self Cite

102

View full text Add to dashboard Cite

show abstract

“…S1, Figure 3) that utilizes site-specifically 2-aminopurine (Ap) labeled RNA [35]. For preQ 1 class-I riboswitches, we focused on the specific aptamer sequence from Thermoanaerobacter tengcongensis (Tte) ; for preQ 1 class-II riboswitches, we used the aptamer sequence from Streptococcus pneumoniae (Spn) (Figure 4A).…”

Section: Resultsmentioning

confidence: 99%

Superior cellular activities of azido- over amino-functionalized ligands for engineered preQ₁riboswitches inE.coli

Neuner¹,

Frener²,

Lusser

et al. 2018

RNA Biology

Self Cite

View full text Add to dashboard Cite

For this study, we utilized class-I and class-II preQ1-sensing riboswitches as model systems to decipher the structure-activity relationship of rationally designed ligand derivatives in vitro and in vivo. We found that synthetic preQ1 ligands with amino-modified side chains that protrude from the ligand-encapsulating binding pocket, and thereby potentially interact with the phosphate backbone in their protonated form, retain or even increase binding affinity for the riboswitches in vitro. They, however, led to significantly lower riboswitch activities in a reporter system in vivo in E. coli. Importantly, when we substituted the amino- by azido-modified side chains, the cellular activities of the ligands were restored for the class-I conditional gene expression system and even improved for the class-II counterpart. Kinetic analysis of ligand binding in vitro revealed enhanced on-rates for amino-modified derivatives while they were attenuated for azido-modified variants. This shows that neither high affinities nor fast on-rates are necessarily translated into efficient cellular activities. Taken together, our comprehensive study interconnects in vitro kinetics and in vitro thermodynamics of RNA-ligand binding with the ligands’ in vivo performance and thereby encourages azido- rather than amino-functionalized design for enhanced cellular activity.

show abstract

“…Their results on E. coli thiM TPP aptamer indicated that the P1 switch helix forms a primarily folded structure in the presence of Mg 2+ alone. Also, P1 and other regions around TPP-binding site exhibit an unexpected degree of plasticity which probably results in facilitating the entry and exit of the TPP ligand and influence on gene regulation (Haller et al, 2013).…”

Section: Discussionmentioning

confidence: 99%

TPP riboswitch characterization in Alishewanella tabrizica and Alishewanella aestuarii and comparison with other TPP riboswitches

Aghdam

Sinn

Tarhriz

et al. 2017

Microbiological Research

View full text Add to dashboard Cite

a b s t r a c tRiboswitches are located in non-coding areas of mRNAs and act as sensors of cellular small molecules, regulating gene expression in response to ligand binding. The TPP riboswitch is the most widespread riboswitch occurring in all three domains of life. However, it has been rarely characterized in environmental bacteria other than Escherichia coli and Bacillus subtilis. In this study, TPP riboswitches located in the 5 UTR of thiC operon from Alishewanella tabrizica and Alishewanella aestuarii were identified and characterized. Moreover, affinity analysis of TPP binding to the TPP aptamer domains originated from A. tabrizica, A. aestuarii, E.coli, and B. subtilis were studied and compared using In-line probing and Surface Plasmon Resonance (SPR). TPP binding to the studied RNAs from A. tabrizica and A. aestuarii caused distinctive changes of the In-line cleavage pattern, demonstrating them as functional TPP riboswitches. With dissociation constant of 2-4 nM (depending on the method utilized), the affinity of TPP binding was highest in A. tabrizica, followed by the motifs sourced from A. aestuarii, E. coli, and B. subtilis. The observed variation in their TPP-binding affinity might be associated with adaptation to the different environments of the studied bacteria.

show abstract

Folding and ligand recognition of the TPP riboswitch aptamer at single-molecule resolution

Cited by 115 publications

References 49 publications

Tuning a riboswitch response through structural extension of a pseudoknot

Tuning a riboswitch response through structural extension of a pseudoknot

Superior cellular activities of azido- over amino-functionalized ligands for engineered preQ₁riboswitches inE.coli

TPP riboswitch characterization in Alishewanella tabrizica and Alishewanella aestuarii and comparison with other TPP riboswitches

Contact Info

Product

Resources

About

Folding and ligand recognition of the TPP riboswitch aptamer at single-molecule resolution

Cited by 115 publications

References 49 publications

Tuning a riboswitch response through structural extension of a pseudoknot

Tuning a riboswitch response through structural extension of a pseudoknot

Superior cellular activities of azido- over amino-functionalized ligands for engineered preQ1riboswitches inE.coli

TPP riboswitch characterization in Alishewanella tabrizica and Alishewanella aestuarii and comparison with other TPP riboswitches

Contact Info

Product

Resources

About

Superior cellular activities of azido- over amino-functionalized ligands for engineered preQ₁riboswitches inE.coli