Thermodynamics and folding kinetics analysis of the SH3 domain form discrete molecular dynamics

We perform a detailed analysis of the thermodynamics and folding kinetics of the SH3 domain fold with discrete molecular dynamic simulations. We propose a protein model that reproduces some of the experimentally observed thermodynamic and folding kinetic properties of proteins. Specifically, we use our model to study the transition state ensemble of the SH3 fold family of proteins, a set of unstable conformations that fold to the protein native state with probability 1/2. We analyze the participation of each secondary structure element formed at the transition state ensemble. We also identify the folding nucleus of the SH3 fold and test extensively its importance for folding kinetics. We predict that a set of amino acid contacts between the RT-loop and the distal hairpin are the critical folding nucleus of the SH3 fold and propose a hypothesis that explains this result.     

ProbKnot: Fast prediction of RNA secondary structure including pseudoknots

It is a significant challenge to predict RNA secondary structures including pseudoknots. Here, a new algorithm capable of predicting pseudoknots of any topology, ProbKnot, is reported. ProbKnot assembles maximum expected accuracy structures from computed base-pairing probabilities in O(N²) time, where N is the length of the sequence. The performance of ProbKnot was measured by comparing predicted structures with known structures for a large database of RNA sequences with fewer than 700 nucleotides. The percentage of known pairs correctly predicted was 69.3%. Additionally, the percentage of predicted pairs in the known structure was 61.3%. This performance is the highest of four tested algorithms that are capable of pseudoknot prediction. The program is available for download at: http://rna.urmc.rochester.edu/RNAstructure.html.     

Computationally reconstructing cotranscriptional RNA folding from experimental data reveals rearrangement of non-native folding intermediates

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Evaluation of current techniques for Ab initio protein structure prediction

The results of a protein structure prediction contest are reviewed. Twelve different groups entered predictions on 14 proteins of known sequence whose structures had been determined but not yet disseminated to the scientific community. Thus, these represent true tests of the current state of structure prediction methodologies. From this work, it is clear that accurate tertiary structure prediction is not yet possible. However, protein fold and motif prediction are possible when the motif is recognizably similar to another known structure. Internal symmetry and the information inherent in an aligned family of homologous sequences facilitate predictive efforts. Novel folds remain a major challenge for prediction efforts. © 1995 Wiley-Liss, Inc.     

A coarse-grained Langevin molecular dynamics approach to de novo protein structure prediction

De novo prediction of protein structures, the prediction of structures from amino acid sequences which are not similar to those of hitherto resolved structures, has been one of the major challenges in molecular biophysics. In this paper, we develop a new method of de novo prediction, which combines the fragment assembly method and the simulation of physical folding process: structures which have consistently assembled fragments are dynamically searched by Langevin molecular dynamics of conformational change. The benchmarking test shows that the prediction is improved when the candidate structures are cross-checked by an empirically derived score function.     

Ab initio folding simulation of the Trp-cage mini-protein approaches NMR resolution

Here, we report a 100 ns molecular dynamics simulation of the folding process of a recently designed autonomous-folding mini-protein designated as tc5b with a new AMBER force field parameter set developed based on condensed-phase quantum mechanical calculations and a Generalized Born continuum solvent model. Starting from its fully extended conformation, our simulation has produced a final structure resembling that of NMR native structure to within 1A main-chain root mean square deviation. Remarkably, the simulated structure stayed in the native state for most part of the simulation after it reached the state. Of greater significance is that our simulation has not only reached the correct main-chain conformation, but also a very high degree of accuracy in side-chain packing conformation. This feat has traditionally been a challenge for ab initio simulation studies. In addition to characterization of the trajectory, comparison of our results to experimental data is also presented. Analysis of the trajectory suggests that the rate-limiting step of folding of this mini-protein is the packing of the Trp side-chain.     

RNA-Puzzles: a CASP-like evaluation of RNA three-dimensional structure prediction

We report the results of a first, collective, blind experiment in RNA three-dimensional (3D) structure prediction, encompassing three prediction puzzles. The goals are to assess the leading edge of RNA structure prediction techniques; compare existing methods and tools; and evaluate their relative strengths, weaknesses, and limitations in terms of sequence length and structural complexity. The results should give potential users insight into the suitability of available methods for different applications and facilitate efforts in the RNA structure prediction community in ongoing efforts to improve prediction tools. We also report the creation of an automated evaluation pipeline to facilitate the analysis of future RNA structure prediction exercises.     

Molecular dynamics characterisations of the Trp-cage folding mechanisms: in the absence and presence of water solvents

Protein folding is an important and yet challenging topic in current molecular biology. In this work, the folding dynamics and mechanisms of the Trp-cage mini-protein were studied with molecular dynamics simulations, in the absence and presence of water solvents. The important intermediates during the Trp-cage folding were determined by gradually decreasing the simulation temperature. The folding transition temperature was identified to be approximately 400 K, and the folding pathway was decomposed into six steps: U ? I ₁ ? I ₂ ? I ₃ ? I ₄ ? F ₁ ? F ₂, where U, I and F represent the unfolded, intermediate and folded states, respectively. The finding that the two helical subunits are successively formed is consistent with the experimental observations, and the Asp9/Arg16 salt bridge forms at the final stage and does not play a significant role during folding kinetics. The presence of water solvents induces hydrophobic collapse as the whole cage comparatively closes. Within aqueous solutions, the Trp-cage folding begins to contract into the meta-stable state, and by traversing the transition state it arrives at the native-like structure, which resembles the experimental structure closely.     

Ab initio molecular dynamics study of the hydration of the formohydroxamate anion

We apply ab initio molecular dynamics (AIMD) to study the hydration structures and electronic properties of the formohydroxamate anion in liquid water. We consider the cis- nitrogen-deprotonated, cis- oxygen-deprotonated, and trans- oxygen-deprotonated formohydroxamate tautomers. They form an average of 6.3, 6.9, and 6.0 hydrogen bonds with water molecules, respectively. The predicted pair correlation functions and time dependence of the hydration numbers suggest that water is highly structured around the nominally negatively charged oxime oxygen in O-deprotonated tautomers but significantly less so around the nitrogen atom in the N-deprotonated species. Wannier function analysis suggests that, in the O-deprotonated anions, the negative charge is concentrated on the oxime oxygen, while in the N-deprotonated case, it is partially delocalized between the nitrogen and the adjoining oxime oxygen atom.     

Secondary structure prediction and folding of globular protein: refolding of ferredoxin

The physicochemical mechanism of protein folding has been elucidated by the island model, describing a growth type of folding. The folding pathway is closely related with nucleation on the polypeptide chain and thus the formation of small local structures or secondary structures at the earliest stage of folding is essential to all following steps. The island model is applicable to any protein, but a high precision of secondary structure prediction is indispensable to folding simulation. The secondary structures formed at the earliest stage of folding are supposed to be of standard form, but they are usually deformed during the folding process, especially at the last stage, although the degree of deformation is different for each protein. Ferredoxin is an example of a protein having this property. According to X-ray investigation (1FDX), ferredoxin is not supposed to have secondary structures. However, if we assumed that in ferredoxin all the residues are in a coil state, we could not attain the correct structure similar to the native one. Further, we found that some parts of the chain are not flexible, suggesting the presence of secondary structures, in agreement with the recent PDB data (1DUR). Assuming standard secondary structures (-helices and -strands) at the nonflexible parts at the early stage of folding, and deforming these at the final stage, a structure similar to the native one was obtained. Another peculiarity of ferredoxin is the absence of disulfide bonds, in spite of its having eight cysteines. The reason cysteines do not form disulfide bonds became clear by applying the lampshade criterion, but more importantly, the two groups of cysteines are ready to make iron complexes, respectively, at a rather later stage of folding. The reason for poor prediction accuracy of secondary structure with conventional methods is discussed.     

Improved prediction of RNA tertiary structure with insights into native state dynamics

The importance of RNA tertiary structure is evident from the growing number of published high resolution NMR and X-ray crystallographic structures of RNA molecules. These structures provide insights into function and create a knowledge base that is leveraged by programs such as Assemble, ModeRNA, RNABuilder, NAST, FARNA, Mc-Sym, RNA2D3D, and iFoldRNA for tertiary structure prediction and design. While these methods sample native-like RNA structures during simulations, all struggle to capture the native RNA conformation after scoring. We propose RSIM, an improved RNA fragment assembly method that preserves RNA global secondary structure while sampling conformations. This approach enhances the quality of predicted RNA tertiary structure, provides insights into the native state dynamics, and generates a powerful visualization of the RNA conformational space. RSIM is available for download from http://www.github.com/jpbida/rsim.     

RNA-Puzzles Round II: assessment of RNA structure prediction programs applied to three large RNA structures

This paper is a report of a second round of RNA-Puzzles, a collective and blind experiment in three-dimensional (3D) RNA structure prediction. Three puzzles, Puzzles 5, 6, and 10, represented sequences of three large RNA structures with limited or no homology with previously solved RNA molecules. A lariat-capping ribozyme, as well as riboswitches complexed to adenosylcobalamin and tRNA, were predicted by seven groups using RNAComposer, ModeRNA/SimRNA, Vfold, Rosetta, DMD, MC-Fold, 3dRNA, and AMBER refinement. Some groups derived models using data from state-of-the-art chemical-mapping methods (SHAPE, DMS, CMCT, and mutate-and-map). The comparisons between the predictions and the three subsequently released crystallographic structures, solved at diffraction resolutions of 2.5–3.2 Å, were carried out automatically using various sets of quality indicators. The comparisons clearly demonstrate the state of present-day de novo prediction abilities as well as the limitations of these state-of-the-art methods. All of the best prediction models have similar topologies to the native structures, which suggests that computational methods for RNA structure prediction can already provide useful structural information for biological problems. However, the prediction accuracy for non-Watson–Crick interactions, key to proper folding of RNAs, is low and some predicted models had high Clash Scores. These two difficulties point to some of the continuing bottlenecks in RNA structure prediction. All submitted models are available for download at http://ahsoka.u-strasbg.fr/rnapuzzles/.     

Relative flexibility of DNA and RNA: a molecular dynamics study

State of the art molecular dynamics simulations are used to study the structure, dynamics, molecular interaction properties and flexibility of DNA and RNA duplexes in aqueous solution. Special attention is paid to the deformability of both types of structures, revisiting concepts on the relative flexibility of DNA and RNA duplexes. Our simulations strongly suggest that the concepts of flexibility, rigidity and deformability are much more complex than usually believed, and that it is not always true that DNA is more flexible than RNA.     

Dimerization of the p53 oligomerization domain: identification of a folding nucleus by molecular dynamics simulations

Dimerization of the p53 oligomerization domain involves coupled folding and binding of monomers. To examine the dimerization, we have performed molecular dynamics (MD) simulations of dimer folding from the rate-limiting transition state ensemble (TSE). Among 799 putative transition state structures that were selected from a large ensemble of high-temperature unfolding trajectories, 129 were identified as members of the TSE via calculation of a 50% transmission coefficient from at least 20 room-temperature simulations. This study is the first to examine the refolding of a protein dimer using MD simulations in explicit water, revealing a folding nucleus for dimerization. Our atomistic simulations are consistent with experiment and offer insight that was previously unobtainable.     

Secondary structure prediction for aligned RNA sequences

Most functional RNA molecules have characteristic secondary structures that are highly conserved in evolution. Here we present a method for computing the consensus structure of a set aligned RNA sequences taking into account both thermodynamic stability and sequence covariation. Comparison with phylogenetic structures of rRNAs shows that a reliability of prediction of more than 80% is achieved for only five related sequences. As an application we show that the Early Noduline mRNA contains significant secondary structure that is supported by sequence covariation.     

Molecular mechanism for preQ1-II riboswitch function revealed by molecular dynamics

Riboswitches are RNA molecules that regulate gene expression using conformational change, affected by binding of small molecule ligands. A crystal structure of a ligand-bound class II preQ₁ riboswitch has been determined in a previous structural study. To gain insight into the dynamics of this riboswitch in solution, eight total molecular dynamic simulations, four with and four without ligand, were performed using the Amber force field. In the presence of ligand, all four of the simulations demonstrated rearranged base pairs at the 3′ end, consistent with expected base-pairing from comparative sequence analysis in a prior bioinformatic analysis; this suggests the pairing in this region was altered by crystallization. Additionally, in the absence of ligand, three of the simulations demonstrated similar changes in base-pairing at the ligand binding site. Significantly, although most of the riboswitch architecture remained intact in the respective trajectories, the P3 stem was destabilized in the ligand-free simulations in a way that exposed the Shine–Dalgarno sequence. This work illustrates how destabilization of two major groove base triples can influence a nearby H-type pseudoknot and provides a mechanism for control of gene expression by a fold that is frequently found in bacterial riboswitches.     

Understanding the folding and stability of a designed WW domain protein with replica exchange molecular dynamics simulations

WW domain proteins are usually regarded as simple models for understanding the folding mechanism of β-sheet. CC45 is an artificial protein that is capable of folding into the same structure as WW domain. In this article, the replica exchange molecular dynamics simulations are performed to investigate the folding mechanism of CC45. The analysis of thermal stability shows that β-hairpin 1 is more stable than β-hairpin 2 during the unfolding process. Free energy analysis shows that the unfolding of this protein substantially proceeds through solvating the smaller β-hairpin 2, followed by the unfolding of β-hairpin 1. We further propose the unfolding process of CC45 and the folding mechanism of two β-hairpins. These results are similar to the previous folding studies of formin binding protein 28 (FBP28). Compared with FBP28, it is found that CC45 has more aromatic residues in N-terminal loop, and these residues contact with C-terminal loop to form the outer hydrophobic core, which increases the stability of CC45. Knowledge about the stability and folding behaviour of CC45 may help in understanding the folding mechanisms of the β-sheet and in designing new WW domains.