Structural Insights into Riboswitch Control of the Biosynthesis of Queuosine, a Modified Nucleotide Found in the Anticodon of tRNA

Kang, Mijeong; Peterson, R. R.; Feigon, Juli

doi:10.1016/j.molcel.2010.08.007

Cited by 34 publications

(64 citation statements)

References 0 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…Structures of transcriptional (Bacillus subtilis queC; Bsu) (33,34) and translational (Thermoanaerobacter tengcongensis; Tte) (35) preQ 1 -I riboswitches in complex with preQ 1 have been reported previously. Both preQ 1 -I and preQ 1 -II riboswitches form H-type pseudoknots when bound to preQ 1 , and each has interactions along three edges of the preQ 1 using one nucleobase each in loop 1, loop 2, and loop 3.…”

Section: Resultsmentioning

confidence: 83%

“…preQ 1 was synthesized as described previously (33) and was dissolved in filtered (0.2-μM filter; Nalgen) nanopure water to make a 50 mM stock solution. Unlabeled, base-specifically and uniformly 13 C, 15 Nlabeled, and partially deuterated (deuterated at the ribose-3′, 4′, 5′, 5′′ positions of all A, U, G, and C and the 5 position of pyrimidines; Cambridge Isotope Laboratories) Spn preQ 1 -II riboswitch WT and mutant samples were prepared by in vitro transcription using T7 polymerase (P266L mutant) with synthetic DNA templates and purified by denaturing polyacrylamide gel electrophoresis.…”

Section: Methodsmentioning

confidence: 99%

“…A suite of 2D-filtered/ edited labeled proton NOESY (F2f, F1fF2e, F1fF2f, and F1eF2e) experiments on base-specifically 13 C, 15 N-labeled RNA samples were used to resolve ambiguous assignments in overlapped regions (69). The assignments of preQ 1 and interactions in the binding pocket were identified as described previously (33). Single-bond C-H and N-H RDCs were measured using 2D 1 H-13 C S3CT-HSQC and standard 1 H-15 N HSQC experiments as described previously (70), and are listed in Table S3.…”

Section: Methodsmentioning

confidence: 99%

“…preQ 1 -I riboswitches have among the smallest known minimal aptamers, at 34 nt (26), but preQ 1 -II riboswitch aptamers are much larger, at ∼60 nt (27). High-resolution X-ray crystal and solution NMR structures have revealed that preQ 1 -I aptamers form H-type pseudoknots with two stems and three loops (33)(34)(35)(36). Subsequent studies detailing preQ 1 -I riboswitch dynamics and folding pathway have shown that divalent cations bind loop 1 and stabilize the bound conformation (21, 37).…”

mentioning

confidence: 99%

See 3 more Smart Citations

Structural determinants for ligand capture by a class II preQ ₁ riboswitch

Kang

Eichhorn²,

Feigon

2014

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

Self Cite

View full text Add to dashboard Cite

Prequeuosine (preQ 1 ) riboswitches are RNA regulatory elements located in the 5′ UTR of genes involved in the biosynthesis and transport of preQ 1 , a precursor of the modified base queuosine universally found in four tRNAs. The preQ 1 class II (preQ 1 -II) riboswitch regulates preQ 1 biosynthesis at the translational level. We present the solution NMR structure and conformational dynamics of the 59 nucleotide Streptococcus pneumoniae preQ 1 -II riboswitch bound to preQ 1 . Unlike in the preQ 1 class I (preQ 1 -I) riboswitch, divalent cations are required for high-affinity binding. The solution structure is an unusual H-type pseudoknot featuring a P4 hairpin embedded in loop 3, which forms a three-way junction with the other two stems. 13 C relaxation and residual dipolar coupling experiments revealed interhelical flexibility of P4. We found that the P4 helix and flanking adenine residues play crucial and unexpected roles in controlling pseudoknot formation and, in turn, sequestering the Shine-Dalgarno sequence. Aided by divalent cations, P4 is poised to act as a "screw cap" on preQ 1 recognition to block ligand exit and stabilize the binding pocket.Comparison of preQ 1 -I and preQ 1 -II riboswitch structures reveals that whereas both form H-type pseudoknots and recognize preQ 1 using one A, C, or U nucleotide from each of three loops, these nucleotides interact with preQ 1 differently, with preQ 1 inserting into different grooves. Our studies show that the preQ 1 -II riboswitch uses an unusual mechanism to harness exquisite control over queuosine metabolism.R iboswitches are functional noncoding RNA elements often found at the 5′ end of genes that bind to effector molecules, usually metabolites, to attenuate gene expression (1-8). They generally contain two modular elements, an aptamer that binds directly to a cognate ligand, followed by an expression platform that carries out the regulatory function. Ligand binding induces a conformational rearrangement in the aptamer domain, which is transduced to the expression platform, turning gene expression on or off at the transcription or translation level.Structural studies of diverse riboswitches bound to their cognate ligands have revealed that they have evolved various strategies for high-affinity and specific binding between RNA receptors and ligands. In general, the aptamer domain enfolds the ligand, forming a highly complex 3D conformation using a host of tertiary interactions, including base triples, kink turns, ribose zippers, A-minor motifs, loop-loop interactions, and pseudoknots (9-11). Some riboswitches require divalent cations for ligand binding (9, 12-16), whereas in others divalent cations stabilize the bound state (17-21). Some ligands, including S-adenosyl methionine (22), S-adenosyl homocysteine (22, 23), cyclic di-GMP (24, 25), and prequeuosine (preQ 1 ) (26, 27), have more than one class of riboswitch, with each class adopting a different 3D fold to achieve recognition of the same ligand.Two classes of riboswitches that bind preQ 1 , a precursor t...

show abstract

Section: Resultsmentioning

confidence: 83%

Section: Methodsmentioning

confidence: 99%

Section: Methodsmentioning

confidence: 99%

mentioning

confidence: 99%

See 2 more Smart Citations

Structural determinants for ligand capture by a class II preQ ₁ riboswitch

Kang

Eichhorn²,

Feigon

2014

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

Self Cite

View full text Add to dashboard Cite

show abstract

“…The preQ 1 -I, preQ 1 -II, S-adenosyl-L-methionine-II (SAM-II), and fluoride riboswitches are representative of this organizational strategy, and their analysis has contributed to a renaissance in our understanding of regulatory pseudoknot structure and dynamics (8)(9)(10)(11)(12)(13)(14)(15)(16)(17)(18). By contrast, pseudoknotted aptamers that do not integrate their expression platforms are less common, and this added complexity can encumber efforts to elucidate how ligand-induced conformational changes regulate gene expression.…”

mentioning

confidence: 99%

Structural analysis of a class III preQ ₁ riboswitch reveals an aptamer distant from a ribosome-binding site regulated by fast dynamics

Liberman

Suddala

Aytenfisu

et al. 2015

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

View full text Add to dashboard Cite

PreQ 1 -III riboswitches are newly identified RNA elements that control bacterial genes in response to preQ 1 (7-aminomethyl-7-deazaguanine), a precursor to the essential hypermodified tRNA base queuosine. Although numerous riboswitches fold as H-type or HL out -type pseudoknots that integrate ligand-binding and regulatory sequences within a single folded domain, the preQ 1 -III riboswitch aptamer forms a HL out -type pseudoknot that does not appear to incorporate its ribosome-binding site (RBS). To understand how this unusual organization confers function, we determined the crystal structure of the class III preQ 1 riboswitch from Faecalibacterium prausnitzii at 2.75 Å resolution. PreQ 1 binds tightly (K D,app 6.5 ± 0.5 nM) between helices P1 and P2 of a three-way helical junction wherein the third helix, P4, projects orthogonally from the ligand-binding pocket, exposing its stem-loop to base pair with the 3′ RBS. Biochemical analysis, computational modeling, and single-molecule FRET imaging demonstrated that preQ 1 enhances P4 reorientation toward P1-P2, promoting a partially nested, H-type pseudoknot in which the RBS undergoes rapid docking (k dock ∼0.6 s −1 ) and undocking (k undock ∼1.1 s −1 ). Discovery of such dynamic conformational switching provides insight into how a riboswitch with bipartite architecture uses dynamics to modulate expression platform accessibility, thus expanding the known repertoire of gene control strategies used by regulatory RNAs. preQ 1 riboswitch | gene regulation | crystal structure | single-molecule FRET | molecular dynamics R iboswitches are structured RNA motifs that sense the cellular levels of small molecules to provide feedback regulation of genes (1). Although present in all domains of life, they are prominent in bacteria where they typically reside in the 5′-leader sequences of mRNA (2). Broad interest in riboswitches originates from the discovery that they can be targeted by antimicrobials (3-5), and the observation that they use complex scaffolds to achieve gene regulation without the need for protein partners. In the latter respect, riboswitches typically exhibit bipartite sequence organization comprising a conserved aptamer linked to a downstream expression platform (2). Aptamer binding to a cognate effector can induce conformational changes that alter the accessibility of expression platform sequences, such as those required for transcriptional read-through, or hybridization to the 16S rRNA as a preface to translation (2, 6).Numerous riboswitches fold as pseudoknots that conform to the H-type or closely related HL out -type topology, which have emerged as the most efficient RNA scaffolds to integrate aptamer and expression platform sequences (7). The preQ 1 -I, preQ 1 -II, S-adenosyl-L-methionine-II (SAM-II), and fluoride riboswitches are representative of this organizational strategy, and their analysis has contributed to a renaissance in our understanding of regulatory pseudoknot structure and dynamics (8-18). By contrast, pseudoknotted aptamers that do not integr...

show abstract

Riboswitch structure in the ligand‐free state

Liberman

Wedekind

2011

WIREs RNA

View full text Add to dashboard Cite

Molecular investigations of riboswitches bound to small-molecule effectors have produced a wealth of information on how these molecules achieve high affinity and specificity for a target ligand. X-ray crystal structures have been determined for the ligand-free state for representatives of the preQ1-I, SAM-I, lysine, and glycine aptamer classes. These structures in conjunction with complimentary techniques, such as in-line probing, NMR spectroscopy, Förster resonance energy transfer, small-angle scattering, and computational simulations, have demonstrated that riboswitches adopt multiple conformations in the absence of ligand. Despite a number of investigations that support ligand-dependent folding, mounting evidence suggests that free-state riboswitches interact with their effectors in sub-populations of largely pre-folded states as embodied by the principle of conformational selection, which has been documented extensively for protein-mediated ligand interactions. Fundamental riboswitch investigations of the bound and free states have advanced our understanding of RNA folding, ligand recognition, and how these factors culminate in communication between an aptamer and its expression platform. An understanding of these topics is essential to comprehend riboswitch gene regulation at the molecular level, which has already provided a basis to understand the mechanism of action of natural antimicrobials.

show abstract

Structural Insights into Riboswitch Control of the Biosynthesis of Queuosine, a Modified Nucleotide Found in the Anticodon of tRNA

Cited by 34 publications

References 0 publications

Structural determinants for ligand capture by a class II preQ ₁ riboswitch

Structural determinants for ligand capture by a class II preQ ₁ riboswitch

Structural analysis of a class III preQ ₁ riboswitch reveals an aptamer distant from a ribosome-binding site regulated by fast dynamics

Riboswitch structure in the ligand‐free state

Contact Info

Product

Resources

About

Structural Insights into Riboswitch Control of the Biosynthesis of Queuosine, a Modified Nucleotide Found in the Anticodon of tRNA

Cited by 34 publications

References 0 publications

Structural determinants for ligand capture by a class II preQ 1 riboswitch

Structural determinants for ligand capture by a class II preQ 1 riboswitch

Structural analysis of a class III preQ 1 riboswitch reveals an aptamer distant from a ribosome-binding site regulated by fast dynamics

Riboswitch structure in the ligand‐free state

Contact Info

Product

Resources

About

Structural determinants for ligand capture by a class II preQ ₁ riboswitch

Structural determinants for ligand capture by a class II preQ ₁ riboswitch

Structural analysis of a class III preQ ₁ riboswitch reveals an aptamer distant from a ribosome-binding site regulated by fast dynamics