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. 2005 May 10;102(19):6789-94.
doi: 10.1073/pnas.0408314102. Epub 2005 Mar 4.

Mechanical unfolding of RNA hairpins

Changbong Hyeon  1 , D Thirumalai
Affiliations

Mechanical unfolding of RNA hairpins

Changbong Hyeon et al. Proc Natl Acad Sci U S A. .

Abstract

Mechanical unfolding trajectories, generated by applying constant force in optical-tweezer experiments, show that RNA hairpins and the P5abc subdomain of the group I intron unfold reversibly. We use coarse-grained Go-like models for RNA hairpins to explore forced unfolding over a broad range of temperatures. A number of predictions that are amenable to experimental tests are made. At the critical force, the hairpin jumps between folded and unfolded conformations without populating any discernible intermediates. The phase diagram in the force-temperature (f, T) plane shows that the hairpin unfolds by an all-or-none process. The cooperativity of the unfolding transition increases dramatically at low temperatures. Free energy of stability, obtained from time averages of mechanical unfolding trajectories, coincides with ensemble averages, which establishes ergodicity. The hopping time between the native basin of attraction (NBA) and the unfolded basin increases dramatically along the phase boundary. Thermal unfolding is stochastic, whereas mechanical unfolding occurs in "quantized steps" with great variations in the step lengths. Refolding times, upon force quench, from stretched states to the NBA are at least an order of magnitude greater than folding times by temperature quench. Upon force quench from stretched states, the NBA is reached in at least three stages. In the initial stages, the mean end-to-end distance decreases nearly continuously, and there is a sudden transition to the NBA only in the last stage. Because of the generality of the results, we propose that similar behavior should be observed in force quench refolding of proteins.

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Figures

Fig. 1.
Fig. 1.
Coarse-grained representation of RNA. (A) Coarse-grained representation of a nucleotide by using three sites; namely, phosphate (P), sugar (S), and base (B) are given. (B) The secondary structure of the 22-nt P5GA hairpin in which the bonds formed between base pairs are labeled 1–9. The PDB structure (13) and the corresponding structure using the coarse grained model are shown on the right.
Fig. 2.
Fig. 2.
Phase diagram for the P5GA hairpin. (A) Diagram of states obtained by using the fraction of native contacts as the order parameter. The values of the thermal average of the fraction of native contacts, 〈Q〉, are color-coded as indicated on the scale shown on the right. The dashed line is a fit by using Eq. 6 to the locus of points in the (f, T) plane that separates the folded hairpin from the unfolded states. (B) Plot of the phase diagram in the (f, T) plane by using the mean end-to-end distance 〈R〉 as the order parameter. Although the diagram of states is qualitatively similar to A, there are quantitative differences in estimates of Tm at f = 0. However, estimates of threshold force values at T < Tm are similar in A and B.
Fig. 3.
Fig. 3.
Hopping transitions at constant f.(A) Time traces of R at various values of constant force at T = 305 K. At f = 4.8 pN < fm ≈ 6 pN, 〈R〉 fluctuates around at low values, which shows that the NBA is preferentially populated (top time trace). As f ∼ fm (third time trace) the hairpin hops between the folded state (low R value) and unfolded states (R ≈ 10 nm). The transitions occur over a short time interval. These time traces are similar to figure 2C in ref. . (B) Logarithm of the equilibrium constant Keq (computed by using the time traces in A) as a function of f. The red line is a fit with log Keq = 10.4 + 1.79 f.(C) Equilibrium free energy profiles F(R) as a function of R at T = 305 K. The colors represent different f values that are shown in Inset. The arrows give the location of the unfolded basin of attraction.
Fig. 4.
Fig. 4.
Folding cooperativity. (A) Dependence of 〈Q(T, f)〉 as a function of f at various temperatures. (B) Values of (Ωfc)T as a function of temperature. (C) Variation of 〈Q(T, f)〉 as a function of T at various values of f.(D) Dimensionless cooperativity measure (ΩTc)f for 0 ≤ f ≤ 20.
Fig. 5.
Fig. 5.
Hopping transitions along the phase boundary. (A) Free-energy profiles F(R) along the phase boundary (Tm, fm) (see Fig. 2). The barrier separating NBA and UBA increases at low Tm and high fm values. (B) Time traces of R obtained by using Brownian dynamics simulations. The values of T and f are 305 K and 6 pN, respectively. The arrows (black, red, and green) indicate the residence times in the NBA for three trajectories.
Fig. 6.
Fig. 6.
Force-induced unfolding and refolding. (A) Time traces of unfolding of P5GA at a constant force f = 42 pN at T = 254 K monitored by the increase in R. The values of Q at different unfolding stages are given for the trajectory in black. (B) Refolding is initiated by a force quench from the initial value f = 90 pN to f = 0. The five time traces show great variations in the relaxation to the hairpin conformation. However, in all trajectories, R decreases in at least three stages that are explicitly labeled for the trajectory in green. The trajectories in A and B are offset, for clarity.
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References

    1. Onoa, B. & Tinoco, I., Jr. (2004) Curr. Opin. Struct. Biol. 14, 374–379. - PubMed
    1. Treiber, D. K. & Williamson, J. R. (2001) Curr. Opin. Struct. Biol. 11, 309–314. - PubMed
    1. Thirumalai, D., Lee, N., Woodson, S. A. & Klimov, D. K. (2001) Annu. Rev. Phys. Chem. 52, 751–762. - PubMed
    1. Koculi, E., Lee, N., Thirumalai, D. & Woodson, S. A. (2004) J. Mol. Biol. 341, 27–36. - PubMed
    1. Russell, R., Zhuang, X., Babcock, H. P., Millett, I. S., Doniach, S., Chu, S. & Herschlag, D. (2002) Proc. Natl. Acad. Sci. USA 99, 155–160. - PMC - PubMed

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