Structural Basis of Cooperative Ligand Binding by the Glycine Riboswitch

Butler, Ethan B.; Xiong, Yong; Wang, Jimin; Strobel, Scott A.

doi:10.1016/j.chembiol.2011.01.013

Cited by 91 publications

(134 citation statements)

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“…Stabilization of a junctional region by ligand binding is a recurrent theme in riboswitch RNAs (16,17). The THF riboswitch joins this common trend except that its ligand binds adjacent to the junction, a feature also observed for the glycine riboswitch (18,19). Projection of the THF-induced modulations (2) (Fig.…”

Section: Resultsmentioning

confidence: 99%

Long-range pseudoknot interactions dictate the regulatory response in the tetrahydrofolate riboswitch

Huang

Ishibe-Murakami

Patel

et al. 2011

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

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Tetrahydrofolate (THF), a biologically active form of the vitamin folate (B 9 ), is an essential cofactor in one-carbon transfer reactions. In bacteria, expression of folate-related genes is controlled by feedback modulation in response to specific binding of THF and related compounds to a riboswitch. Here, we present the X-ray structures of the THF-sensing domain from the Eubacterium siraeum riboswitch in the ligand-bound and unbound states. The structure reveals an "inverted" three-way junctional architecture, most unusual for riboswitches, with the junction located far from the regulatory helix P1 and not directly participating in helix P1 formation. Instead, the three-way junction, stabilized by binding to the ligand, aligns the riboswitch stems for long-range tertiary pseudoknot interactions that contribute to the organization of helix P1 and therefore stipulate the regulatory response of the riboswitch. The pterin moiety of the ligand docks in a semiopen pocket adjacent to the junction, where it forms specific hydrogen bonds with two moderately conserved pyrimidines. The aminobenzoate moiety stacks on a guanine base, whereas the glutamate moiety does not appear to make strong interactions with the RNA. In contrast to other riboswitches, these findings demonstrate that the THF riboswitch uses a limited number of available determinants for ligand recognition. Given that modern antibiotics target folate metabolism, the THF riboswitch structure provides insights on mechanistic aspects of riboswitch function and may help in manipulating THF levels in pathogenic bacteria.RNA structure | vitamin B9 | tetrahydrobiopterin | coenzyme T he tetrahydrofolate (THF) compounds, biologically active reduced derivatives of folate (vitamin B 9 ), are essential cofactors in one-carbon transfer reactions involved in the biosynthesis of many critical molecules. Therefore, maintaining an adequate cellular level of THF is of primary importance to all living beings. In bacteria, THF can be generated from folate transported from the environment using a special transport system (1) or synthesized de novo. Expression of folate transport and synthetic genes is controlled by riboswitches that respond to THF and related compounds (2).Structured mRNA segments termed riboswitches act as both direct sensors of cellular metabolites and effectors of the regulatory response in all three kingdoms of life (3, 4). Typically, specific binding of a cognate metabolite stabilizes the metabolite-bound conformation of the sensing domain of the riboswitch and, through formation of the regulatory helix P1, directs the folding of the adjacent expression platform that carries signals for transcriptional or translational machineries. If the metabolite concentration does not reach a threshold, the riboswitch adopts an alternative conformation, resulting in an opposite effect on gene expression. Riboswitches respond to various types of metabolites; however, the most widespread and abundant riboswitches, presently counting nine classes, are selective to protei...

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Section: Resultsmentioning

confidence: 99%

Long-range pseudoknot interactions dictate the regulatory response in the tetrahydrofolate riboswitch

Huang

Ishibe-Murakami

Patel

et al. 2011

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

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“…For other tandem riboswitches known, an expression platform follows each aptamer domain (Sudarsan et al 2006;Welz and Breaker 2007). Crystallographic structure determination demonstrates that isolated glycine riboswitch aptamer domains adopt the same fold as in tandem (Huang et al 2010;Butler et al 2011). Moreover, both in solution and in crystals, the isolated aptamer domains show a propensity to oligomerize, utilizing what appears to be the same interface as that used by the tandem aptamers when present in a single RNA chain (Huang et al 2010;Butler et al 2011;Erion and Strobel 2011).…”

Section: Introductionmentioning

confidence: 99%

Modulation of quaternary structure and enhancement of ligand binding by the K-turn of tandem glycine riboswitches

Baird

Ferré-D′Amaré

2012

RNA

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Most known glycine riboswitches have two homologous aptamer domains arranged in tandem and separated by a short linker. The two aptamers associate through reciprocal "quaternary" interactions that have been proposed to result in cooperative glycine binding. Recently, the interaptamer linker was found to form helix P0 with a previously unrecognized segment 5′ to the first aptamer domain. P0 was shown to increase glycine affinity, abolish cooperativity, and conform to the K-turn motif consensus. We examine the global thermodynamic and structural role of P0 using isothermal titration calorimetry (ITC) and small-angle X-ray scattering (SAXS), respectively. To evaluate the generality of P0 function, we prepared glycine riboswitch constructs lacking and including P0 from Bacillus subtilis, Fusobacterium nucleatum, and Vibrio cholerae. We find that P0 indeed folds into a K-turn, supports partial pre-folding of all three glycine-free RNAs, and is required for ITC observation of glycine binding under physiologic Mg 2+ concentrations. Except for the unusually small riboswitch from F. nucleatum, the K-turn is needed for maximally compacting the glycine-bound states of the RNAs. Formation of a ribonucleoprotein complex between the B. subtilis or the F. nucleatum RNA constructs and the bacterial K-turn binding protein YbxF promotes additional folding of the free riboswitch, and enhances glycine binding. Consistent with the previously reported loss of cooperativity, P0-containing B. subtilis and V. cholerae tandem aptamers bound no more than one glycine molecule per riboswitch. Our results indicate that the P0 K-turn helps organize the quaternary structure of tandem glycine riboswitches, thereby facilitating ligand binding under physiologic conditions.

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“…3). This glycine riboswitch is conserved across many bacterial species and occurs upstream of genes encoding proteins of the glycine cleavage system, which facilitates use of glycine as an energy source (Butler et al, 2011;Erion & Strobel, 2011;Huang et al, 2010;Kwon & Strobel., 2008;Mandal et al, 2004;Sherman et al, 2012). In Bacillus subtilis, it has been shown that this riboswitch regulates transcription of the downstream gcvT operon (Mandal et al, 2004;Phan & Schumann, 2007).…”

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confidence: 99%

Refinement of the Listeria monocytogenes σB regulon through quantitative proteomic analysis

et al. 2013

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Structural Basis of Cooperative Ligand Binding by the Glycine Riboswitch

Cited by 91 publications

References 25 publications

Long-range pseudoknot interactions dictate the regulatory response in the tetrahydrofolate riboswitch

Long-range pseudoknot interactions dictate the regulatory response in the tetrahydrofolate riboswitch

Modulation of quaternary structure and enhancement of ligand binding by the K-turn of tandem glycine riboswitches

Refinement of the Listeria monocytogenes σB regulon through quantitative proteomic analysis

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