Metal Ion Dependence, Thermodynamics, and Kinetics for Intramolecular Docking of a GAAA Tetraloop and Receptor Connected by a Flexible Linker

Downey, Christopher D.; Fiore, Julie L.; Stoddard, C.D.; Hodak, José H.; Nesbitt, David J.; Pardi, Arthur

doi:10.1021/bi0520941

Cited by 48 publications

(120 citation statements)

References 64 publications

(148 reference statements)

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“…2C; data not shown). These data are consistent with FRET studies that indicate the tetraloop-receptor interaction can form in potassium or cobalt hexammine (Downey et al 2006). …”

Section: Ionic Dependence Of the Tetraloop-receptor Interactionsupporting

confidence: 90%

“…In the presence of 250 mM KCl and 2 mM EDTA, the relative mobility of the GAAA tetraloop-receptor RNA is consistent with formation of the homodimer. These data are also in agreement with recent FRET results of a tethered tetraloop-receptor complex that docks in the presence of potassium (K 1/2 = 200 mM) without divalent cations (Downey et al 2006). Since crystal structures have revealed several ions closely associated with the tetraloop receptor Basu et al 1998;Golden et al 1998; FIGURE 1.…”

Section: Ionic Dependence Of the Tetraloop-receptor Interactionsupporting

confidence: 89%

“…EPR studies have revealed a magnesium dependence for tetraloop docking to the receptor (Qin et al 2005). Recent fluorescence resonance energy transfer (FRET) experiments show a broad metal ion dependence for tetraloop docking, with charge of the ion being the most important factor for determining the concentration of ion required for tetraloop docking (Downey et al 2006).…”

Section: Introductionmentioning

confidence: 99%

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Role of metal ions in the tetraloop–receptor complex as analyzed by NMR

Davis

Foster²,

Tonelli³

et al. 2006

RNA

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Metal ions are critical for the proper folding of RNA, and the GAAA tetraloop-receptor is necessary for the optimal folding and function of many RNAs. We have used NMR to investigate the role of metal ions in the structure of the tetraloop-receptor in solution. The NMR data indicate native tertiary structure is formed under a wide range of ionic conditions. The lack of conformational adaptation in response to very different ionic conditions argues against a structural role for divalent ions. Nuclear Overhauser effects to cobalt hexammine and paramagnetic relaxation enhancement induced by manganese ions were used to determine the NMR structures of the tetraloop receptor in association with metal ions, providing the first atomic-level view of these interactions in the solution state. Five manganese and two cobalt hexammine ions could be localized to the RNA surface. The locations of the associated metal ions are similar, but not identical to, those of previously determined crystal structures. The sites of association are in general agreement with nonlinear Poisson-Boltzmann calculations of the electrostatic surface, emphasizing the general importance of diffusely associated ions in RNA tertiary structure.

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“…2C; data not shown). These data are consistent with FRET studies that indicate the tetraloop-receptor interaction can form in potassium or cobalt hexammine (Downey et al 2006). …”

Section: Ionic Dependence Of the Tetraloop-receptor Interactionsupporting

confidence: 90%

Section: Ionic Dependence Of the Tetraloop-receptor Interactionsupporting

confidence: 89%

Section: Introductionmentioning

confidence: 99%

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Role of metal ions in the tetraloop–receptor complex as analyzed by NMR

Davis

Foster²,

Tonelli³

et al. 2006

RNA

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“…Mg II and Na I ) were added, the DNA collapsed into a more compact structure due to the screening of the negative charges (but still in random coil), also resulting in a shorter end-to-end distance. [23,[35][36][37] Addition of Hg II to this more compact DNA can still induce the hairpin formation as shown in Figure 1B. However, the amount of distance change might be smaller.…”

Section: Resultsmentioning

confidence: 94%

Metal‐Induced Specific and Nonspecific Oligonucleotide Folding Studied by FRET and Related Biophysical and Bioanalytical Implications

Kiy

Jacobi

Liu

2011

Chemistry A European J

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Abstract.Metal induced nucleic acid folding has been extensively studied with ribozymes, DNAzymes, tRNA and riboswitches. These RNA/DNA molecules usually have a high content of double-stranded regions to support a rigid scaffold. On the other hand, such rigid structural features are not available for many in vitro selected or rationally designed DNA aptamers; they adopt flexible random coil structures in the absence of target molecules. Upon target binding, these aptamers adaptively fold into a compact structure with a reduced end-to-end distance, making fluorescence resonance energy transfer (FRET) a popular signaling mechanism. However, non-specific folding induced by mono-or divalent metal ions can also reduce the end-to-end distance and thus lead to false positive results. In this study we used a FRET pair labeled Hg II binding DNA and monitored metal induced folding in the presence of various cations. While non-specific electrostatically mediated folding can be very significant, at each tested salt condition, Hg II induced folding was still observed with a similar sensitivity. We also studied the biophysical meaning of the acceptor/donor fluorescence ratio that allowed us to explain the experimental observations. Potential solutions for this ionic strength problem have been discussed. For example, probes designed to signaling the formation of double-stranded DNA showed a lower dependency on ionic strength.3

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“…Some insight into the free energy landscapes for RNA folding can be obtained from temperature-dependent stopped-flow kinetic studies, which offer the ability to deconstruct free energy barriers (ΔG ‡ ) into enthalpic (ΔH ‡ ) and entropic (−TΔS ‡ ) components. However, with such methods, only the net rate constant (i.e., k total ¼ k fold þ k unfold ) for approach to equilibrium can be observed, which requires strong assumptions (e.g., that k fold ≫ k unfold or k unfold is temperature independent) to permit accurate extraction of transition-state barrier heights (11,(14)(15)(16). Single-molecule fluorescence resonance energy transfer methods (smFRET) avoid such kinetic restrictions by providing both folding and unfolding rate constants under equilibrium conditions, though smFRET transition-states studies of RNA folding are scarce (9,13,17).…”

mentioning

confidence: 99%

Entropic origin of Mg ²⁺ -facilitated RNA folding

Fiore

Holmstrom

Nesbitt

2012

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

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158

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Mg 2þ is essential for the proper folding and function of RNA, though the effect of Mg 2þ concentration on the free energy, enthalpy, and entropy landscapes of RNA folding is unknown. This work exploits temperature-controlled single-molecule FRET methods to address the thermodynamics of RNA folding pathways by probing the intramolecular docking/undocking kinetics of the ubiquitous GAAA tetraloop−receptor tertiary interaction as a function of [Mg 2þ ]. These measurements yield the barrier and standard state enthalpies, entropies, and free energies for an RNA tertiary transition, in particular, revealing the thermodynamic origin of [Mg 2þ ]-facilitated folding. Surprisingly, these studies reveal that increasing [Mg 2þ ] promotes tetraloop-receptor interaction by reducing the entropic barrier (−T ΔS ‡ dock ) and the overall entropic penalty (−T ΔS°d ock ) for docking, with essentially negligible effects on both the activation enthalpy (ΔH ‡ dock ) and overall exothermicity (ΔH°d ock ). These observations contrast with the conventional notion that increasing [Mg 2þ ] facilitates folding by minimizing electrostatic repulsion of opposing RNA helices, which would incorrectly predict a decrease in ΔH ‡ dock and ΔH°d ock with [Mg 2þ ]. Instead we propose that higher [Mg 2þ ] can aid RNA folding by decreasing the entropic penalty of counterion uptake and by reducing disorder of the unfolded conformational ensemble.T he folding of RNA proceeds hierarchically, whereby secondary structure is formed rapidly and subsequent slow helical packing is mediated by tertiary interactions (1, 2). RNA secondary structure prediction from the known thermodynamics is quite reliable (3), though correspondingly accurate prediction of tertiary structure remains a major challenge (1). Static tertiary structure data alone are also not enough to predict RNA functionality, as time-dependent conformational dynamics occur during biochemical processes (4, 5). As a result, one needs the full free energy, enthalpy, and entropy landscapes for folding. A major road block in achieving a predictive understanding of RNA folding landscapes is that they are often "rugged,", i.e., with alternative conformations acting as kinetic traps (6, 7). Moreover, the electrostatic challenge of folding a charged biopolymer highlights the particularly critical role of Mg 2þ and other counterions in the folding process.Characterization of folding transition states-and the role of Mg 2þ in stabilizing transition states-remains a crucial bottleneck for reconciling the kinetics and thermodynamics of RNA folding (8-13). Some insight into the free energy landscapes for RNA folding can be obtained from temperature-dependent stopped-flow kinetic studies, which offer the ability to deconstruct free energy barriers (ΔG ‡ ) into enthalpic (ΔH ‡ ) and entropic (−TΔS ‡ ) components. However, with such methods, only the net rate constant (i.e., k total ¼ k fold þ k unfold ) for approach to equilibrium can be observed, which requires strong assumptions (e.g., that k fold ≫ k unfol...

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Metal Ion Dependence, Thermodynamics, and Kinetics for Intramolecular Docking of a GAAA Tetraloop and Receptor Connected by a Flexible Linker

Cited by 48 publications

References 64 publications

Role of metal ions in the tetraloop–receptor complex as analyzed by NMR

Role of metal ions in the tetraloop–receptor complex as analyzed by NMR

Metal‐Induced Specific and Nonspecific Oligonucleotide Folding Studied by FRET and Related Biophysical and Bioanalytical Implications

Entropic origin of Mg ²⁺ -facilitated RNA folding

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Metal Ion Dependence, Thermodynamics, and Kinetics for Intramolecular Docking of a GAAA Tetraloop and Receptor Connected by a Flexible Linker

Cited by 48 publications

References 64 publications

Role of metal ions in the tetraloop–receptor complex as analyzed by NMR

Role of metal ions in the tetraloop–receptor complex as analyzed by NMR

Metal‐Induced Specific and Nonspecific Oligonucleotide Folding Studied by FRET and Related Biophysical and Bioanalytical Implications

Entropic origin of Mg 2+ -facilitated RNA folding

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Entropic origin of Mg ²⁺ -facilitated RNA folding