The α-helix folds on the millisecond time scale

Clarke, David T.; Doig, Andrew J.; Stapley, Benjamin J.; Jones, Gareth R.

doi:10.1073/pnas.96.13.7232

Cited by 115 publications

(140 citation statements)

References 37 publications

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“…Recently, non-Arrhenius kinetics were found in molecular dynamics simulation by using an atomistic model (32). Experimental evidence is also present for ␣-helical peptides (33,34), a ␤-hairpin peptide (12), CI2 and barnase (35), and lysozyme (36). Given the assumption that the interactions in the DNA hairpin-loop are independent of temperature, the unusual temperature dependence of the opening and closing kinetics might simply be a consequence of the temperature dependence of the accessible configuration space (32).…”

Section: Resultsmentioning

confidence: 96%

Non-Arrhenius kinetics for the loop closure of a DNA hairpin

Wallace

Ying

Balasubramanian

et al. 2001

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

183

299

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DNA hairpin-loop structures fluctuate between different conformations and are generally classified as open or closed (as shown in Fig. 1). They are involved in various biological functions, including gene expression and regulation (5, 6), and more recently they have found use as DNA biosensors (e.g., molecular beacons) (7,8).The open-to-closed transition provides a simple case with which to study the dynamics of intramolecular chain diffusion. Continuing this theme, we developed a method for detecting biopolymer conformational dynamics based on the fluctuations in proximity ratio from FRET (10). By attaching donor and acceptor fluorophores to both ends of a DNA hairpin, the open-to-closed conformational dynamics of the system can be detected at ultra-high sensitivity down to the single-molecule level. By constructing the autocorrelation function of the proximity ratio rather than of the fluorescence intensity, we simplify the extraction of intramolecular kinetics from the correlation function. The use of the ratiometric method should result in a correlation function independent of molecular diffusion (10). This new approach enables us to observe stretched exponential kinetics for the conformational fluctuation in a DNA hairpinloop system (10). A multiple-pathway, two-state model was proposed and used to simulate experimental single-molecule proximity ratio distributions (11).In our previous studies, we concentrated on the time scale of the observed fluctuation and did not fully exploit the amplitude of the autocorrelation function. Here, we report the investigation of temperature and viscosity dependence for the conformational fluctuation of a DNA hairpin-loop. We show that the correlation amplitude is directly related to the equilibrium constant of the open-to-closed transition. By using a two-state model we are able to recover the apparent thermodynamic and kinetic parameters for the hairpin-loop conformational fluctuations under different buffer conditions. The kinetics for the open-to-closed transition of the hairpin-loop appear to show non-Arrhenius behavior akin to that found in peptide ␤-hairpins (12). Materials and MethodsA 40-base oligonucleotide 5Ј-GGGTT-(A) 30 -AACCC-3Ј was chosen as our model DNA hairpin-loop. Donor fluorophore carboxytetramethylrhodamine (TMR) is attached at its 3Ј end via a modified cytosine and a six-carbon linker. Acceptor fluorophore indodicarbocyanine (Cy5) is attached at its 5Ј end via a three-carbon linker. The donor and acceptor form a fluorescence resonance energy transfer pair with a FRET distance (R 0 ) of Ϸ5.3 nm. Both dual-labeled and singly (TMR) labeled oligonucleotides were purchased from Operon Technologies (Alameda, CA) and were HPLC purified. The structure of the fully closed hairpin-loop is illustrated in Fig. 1.The viscosity was varied by adding up to 55% (in mass) of glycerol (molecular biology reagent from Sigma) to the aqueous solution. The precise viscosity of the mixture was calculated by using a polynomial fit to the tabulated viscosity of water͞glycerol m...

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Section: Resultsmentioning

confidence: 96%

Non-Arrhenius kinetics for the loop closure of a DNA hairpin

Wallace

Ying

Balasubramanian

et al. 2001

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

183

299

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“…Another question that is also under debate involves the rate of the nucleation process. The T-jump data of infrared (10), fluorescence (11,12), and Raman (13), as well as results from an NMR experiment (16) all suggest that the nucleation step takes place on a submicrosecond time scale, whereas a stopped-flow CD study (17) indicates that the nucleation process may be much slower, on millisecond time scales.Recently, a new view of protein folding, based on statistical mechanical models of protein-like lattice or off-lattice polymers, has gained popularity in explaining protein folding dynamics, especially inhomogeneous folding kinetics. This new view describes protein folding as parallel, diffusion-like motions of a conformation ensemble on a highly dimensional energy surface, i.e., folding energy landscape, biased toward the native state.…”

mentioning

confidence: 99%

mentioning

confidence: 99%

Helix formation via conformation diffusion search

Huang

Getahun

Zhu

et al. 2002

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

223

342

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The helix-coil transition kinetics of an ␣-helical peptide were investigated by time-resolved infrared spectroscopy coupled with laser-induced temperature-jump initiation method. Specific isotope labeling of the amide carbonyl groups with 13 C at selected residues was used to obtain site-specific information. The relaxation kinetics following a temperature jump, obtained by probing the amide I band of the peptide backbone, exhibit nonexponential behavior and are sensitive to both initial and final temperatures. These data are consistent with a conformation diffusion process on the folding energy landscape, in accord with a recent molecular dynamics simulation study.T he helix-coil transition represents the simplest scenario in protein folding (1-9), yet the details of its kinetics are not understood fully. The classical helix-coil transition theory describes the mechanism of helix formation as a sequence of events, starting from a so-called nucleation step where the first helical hydrogen bond, formed between the amide carbonyl of residue i and the amide hydrogen of residue iϩ4, is generated. The subsequent steps involve helix elongation by adding an extra hydrogen bond at either end of the preexisting helical turns. It has been argued that the nucleation process encounters the largest free energy barrier during the course of helix formation because three residues concomitantly lose their conformational entropy, whereas the propagation steps are energetically favorable because only one residue loses conformational entropy that is balanced by the energy generated from the formation of one extra hydrogen bond. Provided that the nucleation barrier is large enough (compared with those encountered by the propagation steps), a two-state scenario as well as transition-state theory remains effective to explain the dynamics of helix formation. Although recent experiments on the helix-coil transition employing laser-induced temperature-jump (T-jump) method (10-13) have shown that single exponential kinetics, which are characteristic of a two-state system, seem to be adequate to describe the transition between helix-containing and nonhelixcontaining conformations, other studies involving theory and molecular dynamics (MD) simulations have suggested that the helix-coil transition may not follow first-order kinetics (14,15). Another question that is also under debate involves the rate of the nucleation process. The T-jump data of infrared (10), fluorescence (11,12), and Raman (13), as well as results from an NMR experiment (16) all suggest that the nucleation step takes place on a submicrosecond time scale, whereas a stopped-flow CD study (17) indicates that the nucleation process may be much slower, on millisecond time scales.Recently, a new view of protein folding, based on statistical mechanical models of protein-like lattice or off-lattice polymers, has gained popularity in explaining protein folding dynamics, especially inhomogeneous folding kinetics. This new view describes protein folding as parallel, diffusion-like m...

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“…Similarly, in nucleation processes such as α-helix formation or melting, local helix turns can form randomly or sequentially. The first time a complete helix forms (or melts) will be an important ingredient in protein folding models [6]. First passage times also define extinction and fixation in birth-death processes [7,8].…”

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confidence: 99%

First Passage and Cooperativity of Queuing Kinetics

D’Orsogna

Chou

2005

Phys. Rev. Lett.

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We model the kinetics of ligand-receptor systems, where multiple ligands may bind and unbind to the receptor, either randomly or in a specific order. Equilibrium occupation and first occurrence of complete filling of the receptor are determined and compared. At equilibrium, receptors that bind ligands sequentially are more likely to be saturated than those that bind in random order. Surprisingly however, for low cooperativity, the random process first reaches full occupancy faster than the sequential one. This is true except near a critical binding energy where a 'kinetic trap' arises and the random process dramatically slows down when the number of binding sites N ≥ 8. These results demonstrate the subtle interplay between cooperativity and sequentiality for a wide class of kinetic phenomena, including chemical binding, nucleation, and assembly line strategies. [4,5]. For a local bulk ligand concentration, the associated receptors will typically have a fraction of sites filled. The kinetics of queuing in these processes can exhibit diverse and rich behavior. A receptor may need to have a critical number of bound ligands before it can signal the next biochemical step. Thus, it is important to know not only the equilibrium ligand occupancy, but also the mean time to first reach this critical occupancy, as a function of local ligand concentration and binding strength. Similarly, in nucleation processes such as α-helix formation or melting, local helix turns can form randomly or sequentially. The first time a complete helix forms (or melts) will be an important ingredient in protein folding models [6]. First passage times also define extinction and fixation in birth-death processes [7,8]. Equilibrium distributions and first passage times also arise in applications of queuing, where, for example, average computer loads and the first time that demand exceeds capacity should be distinguished [9].In this Letter, we formulate and use a kinetic chain model to highlight subtleties of ligand adsorption and desorption, queuing, and cooperativity. Our model is presented in the language of ligand binding to a single receptor with N active sites of which 0 ≤ n ≤ N are occupied by ligands at any given time. The order of the binding can be imposed in two limiting ways. As shown in Fig. 1, the addition of each successive ligand can influence one other specific site and allow the next ligand to bind to, or unbind from, that site only (a case we will denote by the index α = 0). Alternatively, the allosteric effect (from e.g. a large scale conformational change) can be spread equally to all remaining sites. The next ligand can bind to any one of these remaining open sites (a case we will denote by the index α = 1). Here, all bound ligand molecules are equally likely to spontaneously desorb. We do not consider mixed processes in which the binding order is sequential and the unbinding random, or vice versa. Thus, in our model, ligand binding occurs in a totally sequential manner as in Fig. 1a, or randomly as shown in Fig. 1b. We show...

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The α-helix folds on the millisecond time scale

Cited by 115 publications

References 37 publications

Non-Arrhenius kinetics for the loop closure of a DNA hairpin

Non-Arrhenius kinetics for the loop closure of a DNA hairpin

Helix formation via conformation diffusion search

First Passage and Cooperativity of Queuing Kinetics

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