The Role of Counterion Valence and Size in GAAA Tetraloop–Receptor Docking/Undocking Kinetics

Fiore, Julie L.; Holmstrom, Erik D.; Fiegland, Larry R.; Hodak, José H.; Nesbitt, David J.

doi:10.1016/j.jmb.2012.07.006

Cited by 23 publications

(34 citation statements)

References 105 publications

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“…5(b)). Remarkably, the second-order rate constant along this pathway, ~10 5 M −1 min −1 , is close to that for formation of the same tetraloop-receptor pair in an isolated system (5 × 10 5 M −1 min −1 ) [46–48], consistent with this model (see Supporting Information, Estimation of the rate constant for TL/TLR formation ). Further work will be necessary to test this hypothesis.…”

Section: Discussionsupporting

confidence: 79%

“…While the conformational selection pathway relies on formation of native structure in P5abc and kissing loop P14 formation to nucleate assembly, the induced fit pathway depends on a distinct tertiary module, most likely the TL/TLR contact, which is distant from P5c and not expected to be impacted by the local folding transition of P5abc. Interestingly, a complex in which only the TL/TLR contact is formed would be expected to dissociate rapidly, with a rate constant of at least 200 min −1 [47, 48]. Both of the additional tertiary contacts, the MC/MCR and P14, are most simply expected to require the P5c rearrangement and native structure formation in P5abc.…”

Section: Discussionmentioning

confidence: 99%

See 1 more Smart Citation

RNA Structural Modules Control the Rate and Pathway of RNA Folding and Assembly

Gracia

Xue

Bisaria

et al. 2016

Journal of Molecular Biology

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Structured RNAs fold through multiple pathways, but we have little understanding of the molecular features that dictate folding pathways and determine rates along a given pathway. Here we asked whether folding of a complex RNA can be understood from its structural modules. In a two-piece version of the Tetrahymena group I ribozyme, the separated P5abc subdomain folds to local native secondary and tertiary structure in a linked transition and assembles with the ribozyme core via three tertiary contacts: a kissing loop (P14), a metal core-receptor interaction (MC/MCR), and a tetraloop-receptor interaction (TL/TLR), the first two of which are expected to depend on native P5abc structure from the local transition. Native gel, NMR, and chemical footprinting experiments showed that mutations that destabilize the native P5abc structure slowed assembly up to 100-fold, indicating that P5abc folds first and then assembles with the core by conformational selection. However, rate decreases beyond 100-fold were not observed because an alternative pathway becomes dominant, with non-native P5abc binding the core and then undergoing an induced fit rearrangement. P14 is formed in the rate-limiting step along the conformational selection pathway but after the rate-limiting step along the induced fit pathway. Strikingly, the assembly rate along the conformational selection pathway resembles that for an isolated kissing loop similar to P14, and the rate along the induced fit pathway resembles that for an isolated TL/TLR. Our results indicate substantial modularity in RNA folding and assembly and suggest that these processes can be understood in terms of underlying structural modules.

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Section: Discussionsupporting

confidence: 79%

Section: Discussionmentioning

confidence: 99%

RNA Structural Modules Control the Rate and Pathway of RNA Folding and Assembly

Gracia

Xue

Bisaria

et al. 2016

Journal of Molecular Biology

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“…Motivated by ensemble equilibrium measurements of the T-R FRET construct (Fig. 10), which showed that T(GAAA)-R(11 nt) docking is enabled by many different cations (Downey et al 2006), single-molecule methods were next used to investigate the role of cations in the kinetics of docking/undocking (Fiore et al 2012a). Specifically, the cation-dependent kinetics have been studied as a systematic function of cationic size and charge for a series of monovalent (Na + and K + ), divalent (Mg 2+ and Ca 2+ ), and trivalent [Co(NH 3 ) 6 3+ and spermidine 3+ ] ions.…”

Section: Single-molecule Kinetics and Equilibrium Of Tetraloop-receptmentioning

confidence: 99%

An RNA folding motif: GNRA tetraloop–receptor interactions

Fiore

Nesbitt

2013

Quart. Rev. Biophys.

Self Cite

105

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Nearly two decades after Westhof and Michel first proposed that RNA tetraloops may interact with distal helices, tetraloop–receptor interactions have been recognized as ubiquitous elements of RNA tertiary structure. The unique architecture of GNRA tetraloops (N=any nucleotide, R=purine) enables interaction with a variety of receptors, e.g., helical minor grooves and asymmetric internal loops. The most common example of the latter is the GAAA tetraloop–11 nt tetraloop receptor motif. Biophysical characterization of this motif provided evidence for the modularity of RNA structure, with applications spanning improved crystallization methods to RNA tectonics. In this review, we identify and compare types of GNRA tetraloop–receptor interactions. Then we explore the abundance of structural, kinetic, and thermodynamic information on the frequently occurring and most widely studied GAAA tetraloop–11 nt receptor motif. Studies of this interaction have revealed powerful paradigms for structural assembly of RNA, as well as providing new insights into the roles of cations, transition states and protein chaperones in RNA folding pathways. However, further research will clearly be necessary to characterize other tetraloop–receptor and long-range tertiary binding interactions in detail – an important milestone in the quantitative prediction of free energy landscapes for RNA folding.

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“…Unlike proteins, RNA can and does fold into alternative inactive tertiary structures[6–8], which sometimes can be rescued but sometime lead to degradation of the misfolded molecule. To study RNA tertiary structure, model systems are essential, and especially those that utilize a range of interactions that can be probed for their energetic and kinetic contributions to tertiary structure formation[9,10][11,12] . Here, we introduce a 60 nucleotide fragment from prokaryotic 23S rRNA as a new model system for the mechanics of tertiary structure formation.…”

Section: Introductionmentioning

confidence: 99%

Formation of Tertiary Interactions during rRNA GTPase Center Folding

Rau

Welty

Stump

et al. 2015

Journal of Molecular Biology

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The 60 nucleotide GTPase center (GAC) of 23S rRNA has a phylogenetically conserved secondary structure with two hairpin loops and a 3-way junction. It folds into an intricate tertiary structure upon addition of Mg2+ ions, which is stabilized by the L11 protein in cocrystal structures. Here we monitor the kinetics of its tertiary folding and Mg2+-dependent intermediate states by observation of selected nucleobases that contribute specific interactions to the GAC tertiary structure in the cocrystals. The fluorescent nucleobase 2-aminopurine (2AP) replaced three individual adenines, two of which make long-range stacking interactions, and one that also forms hydrogen bonds. Each site reveals a unique response to Mg2+ addition and temperature, reflecting its environmental change from secondary to tertiary structure. Stopped-flow fluorescence experiments revealed that kinetics of tertiary structure formation upon addition of MgCl2 are also site-specific, with local conformational changes occurring from 5 ms – 4 s, and with global folding from 1- 5 s. Site-specific substitution with 15N-nucleobases allowed observation of stable hydrogen bond formation by NMR experiments. Equilibrium titration experiments indicate that a stable folding intermediate is present at stoichiometric concentrations of Mg2+, and suggest that there are two initial sites of Mg2+ ion association.

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The Role of Counterion Valence and Size in GAAA Tetraloop–Receptor Docking/Undocking Kinetics

Cited by 23 publications

References 105 publications

RNA Structural Modules Control the Rate and Pathway of RNA Folding and Assembly

RNA Structural Modules Control the Rate and Pathway of RNA Folding and Assembly

An RNA folding motif: GNRA tetraloop–receptor interactions

Formation of Tertiary Interactions during rRNA GTPase Center Folding

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