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Sample records for folding problem kinetics

  1. Fast protein folding kinetics

    PubMed Central

    Gelman, Hannah; Gruebele, Martin

    2014-01-01

    Fast folding proteins have been a major focus of computational and experimental study because they are accessible to both techniques: they are small and fast enough to be reasonably simulated with current computational power, but have dynamics slow enough to be observed with specially developed experimental techniques. This coupled study of fast folding proteins has provided insight into the mechanisms which allow some proteins to find their native conformation well less than 1 ms and has uncovered examples of theoretically predicted phenomena such as downhill folding. The study of fast folders also informs our understanding of even “slow” folding processes: fast folders are small, relatively simple protein domains and the principles that govern their folding also govern the folding of more complex systems. This review summarizes the major theoretical and experimental techniques used to study fast folding proteins and provides an overview of the major findings of fast folding research. Finally, we examine the themes that have emerged from studying fast folders and briefly summarize their application to protein folding in general as well as some work that is left to do. PMID:24641816

  2. Kinetic Intermediates in RNA Folding

    NASA Astrophysics Data System (ADS)

    Zarrinkar, Patrick P.; Williamson, James R.

    1994-08-01

    The folding pathways of large, highly structured RNA molecules are largely unexplored. Insight into both the kinetics of folding and the presence of intermediates was provided in a study of the Mg2+-induced folding of the Tetrahymena ribozyme by hybridization of complementary oligodeoxynucleotide probes. This RNA folds via a complex mechanism involving both Mg2+-dependent and Mg2+-independent steps. A hierarchical model for the folding pathway is proposed in which formation of one helical domain (P4-P6) precedes that of a second helical domain (P3-P7). The overall rate-limiting step is formation of P3-P7, and takes place with an observed rate constant of 0.72 ± 0.14 minute-1. The folding mechanism of large RNAs appears similar to that of many multidomain proteins in that formation of independently stable substructures precedes their association into the final conformation.

  3. Problem Solving through Paper Folding

    ERIC Educational Resources Information Center

    Wares, Arsalan

    2014-01-01

    The purpose of this article is to describe a couple of challenging mathematical problems that involve paper folding. These problem-solving tasks can be used to foster geometric and algebraic thinking among students. The context of paper folding makes some of the abstract mathematical ideas involved relatively concrete. When implemented…

  4. Kinetic partitioning mechanism of HDV ribozyme folding

    SciTech Connect

    Chen, Jiawen; Gong, Sha; Wang, Yujie; Zhang, Wenbing

    2014-01-14

    RNA folding kinetics is directly tied to RNA biological functions. We introduce here a new approach for predicting the folding kinetics of RNA secondary structure with pseudoknots. This approach is based on our previous established helix-based method for predicting the folding kinetics of RNA secondary structure. In this approach, the transition rates for an elementary step: (1) formation, (2) disruption of a helix stem, and (3) helix formation with concomitant partial melting of an incompatible helix, are calculated with the free energy landscape. The folding kinetics of the Hepatitis delta virus (HDV) ribozyme and the mutated sequences are studied with this method. The folding pathways are identified by recursive searching the states with high net flux-in(out) population starting from the native state. The theory results are in good agreement with that of the experiments. The results indicate that the bi-phasic folding kinetics for the wt HDV sequence is ascribed to the kinetic partitioning mechanism: Part of the population will quickly fold to the native state along the fast pathway, while another part of the population will fold along the slow pathway, in which the population is trapped in a non-native state. Single mutation not only changes the folding rate but also the folding pathway.

  5. Kinetic partitioning mechanism of HDV ribozyme folding

    NASA Astrophysics Data System (ADS)

    Chen, Jiawen; Gong, Sha; Wang, Yujie; Zhang, Wenbing

    2014-01-01

    RNA folding kinetics is directly tied to RNA biological functions. We introduce here a new approach for predicting the folding kinetics of RNA secondary structure with pseudoknots. This approach is based on our previous established helix-based method for predicting the folding kinetics of RNA secondary structure. In this approach, the transition rates for an elementary step: (1) formation, (2) disruption of a helix stem, and (3) helix formation with concomitant partial melting of an incompatible helix, are calculated with the free energy landscape. The folding kinetics of the Hepatitis delta virus (HDV) ribozyme and the mutated sequences are studied with this method. The folding pathways are identified by recursive searching the states with high net flux-in(out) population starting from the native state. The theory results are in good agreement with that of the experiments. The results indicate that the bi-phasic folding kinetics for the wt HDV sequence is ascribed to the kinetic partitioning mechanism: Part of the population will quickly fold to the native state along the fast pathway, while another part of the population will fold along the slow pathway, in which the population is trapped in a non-native state. Single mutation not only changes the folding rate but also the folding pathway.

  6. Kinetic Analysis of Protein Folding Lattice Models

    NASA Astrophysics Data System (ADS)

    Chen, Hu; Zhou, Xin; Liaw, Chih Young; Koh, Chan Ghee

    Based on two-dimensional square lattice models of proteins, the relation between folding time and temperature is studied by Monte Carlo simulation. The results can be represented by a kinetic model with three states — random coil, molten globule, and native state. The folding process is composed of nonspecific collapse and final searching for the native state. At high temperature, it is easy to escape from local traps in the folding process. With decreasing temperature, because of the trapping in local traps, the final searching speed decreases. Then the folding shows chevron rollover. Through the analysis of the fitted parameters of the kinetic model, it is found that the main difference between the energy landscapes of the HP model and the Go model is that the number of local minima of the Go model is less than that of the HP model.

  7. Kinetics of Peptide Folding in Lipid Membranes

    PubMed Central

    Oh, Kwang-Im; Smith-Dupont, Kathryn B.; Markiewicz, Beatrice N.; Gai, Feng

    2015-01-01

    Despite our extensive understanding of water-soluble protein folding kinetics, much less is known about the folding dynamics and mechanisms of membrane proteins. However, recent studies have shown that for relatively simple systems, such as peptides that form a transmembrane α-helix, helical dimer, or helix-turn-helix, it is possible to assess the kinetics of several important steps, including peptide binding to the membrane from aqueous solution, peptide folding on the membrane surface, helix insertion into the membrane, and helix-helix association inside the membrane. Herein, we provide a brief review of these studies and also suggest new initiation and probing methods that could lead to improved temporal and structural resolution in future experiments. PMID:25808575

  8. Stretched versus compressed exponential kinetics in α-helix folding

    NASA Astrophysics Data System (ADS)

    Hamm, Peter; Helbing, Jan; Bredenbeck, Jens

    2006-03-01

    In a recent paper (J. Bredenbeck, J. Helbing, J.R. Kumita, G.A. Woolley, P. Hamm, α-helix formation in a photoswitchable peptide tracked from picoseconds to microseconds by time resolved IR spectroscopy, Proc. Natl. Acad. Sci USA 102 (2005) 2379), we have investigated the folding of a photo-switchable α-helix with a kinetics that could be fit by a stretched exponential function exp(-( t/ τ) β). The stretching factor β became smaller as the temperature was lowered, a result which has been interpreted in terms of activated diffusion on a rugged energy surface. In the present paper, we discuss under which conditions diffusion problems occur with stretched exponential kinetics ( β < 1) and under which compressed exponential kinetics is obtained ( β > 1). We show that diffusion problems do have a strong tendency to yield stretched exponential kinetics, yet, that there are conditions (strong perturbation from equilibrium, performing the experiment in the folding direction) under which compressed exponential kinetics would be expected instead. We discuss the kinetics on free energy surfaces predicted by simple initiation-propagation models (zipper models) of α-helix folding, as well as by folding funnel models. We show that our recent experiment has been performed under condition for which models with strong downhill driving force, such as the zipper model, would predict compressed, rather than stretched exponential kinetics, in disagreement with the experimental observation. We therefore propose that the free energy surface along a reaction coordinate that governs the folding kinetics must be relatively flat and has a shape similar to a 1D golf course. We discuss how this conclusion can be unified with the thermodynamically well established zipper model by introducing an additional kinetic reaction coordinate.

  9. Fast, Approximate Kinetics of RNA Folding

    PubMed Central

    Senter, Evan

    2015-01-01

    Abstract In this article, we introduce the software suite Hermes, which provides fast, novel algorithms for RNA secondary structure kinetics. Using the fast Fourier transform to efficiently compute the Boltzmann probability that a secondary structure S of a given RNA sequence has base pair distance x (resp. y) from reference structure A (resp. B), Hermes computes the exact kinetics of folding from A to B in this coarse-grained model. In particular, Hermes computes the mean first passage time from the transition probability matrix by using matrix inversion, and also computes the equilibrium time from the rate matrix by using spectral decomposition. Due to the model granularity and the speed of Hermes, it is capable of determining secondary structure refolding kinetics for large RNA sequences, beyond the range of other methods. Comparative benchmarking of Hermes with other methods indicates that Hermes provides refolding kinetics of accuracy suitable for use in the computational design of RNA, an important area of synthetic biology. Source code and documentation for Hermes are available. PMID:25684201

  10. The primary dynamics in protein folding: the earliest kinetic steps.

    NASA Astrophysics Data System (ADS)

    Callender, Robert

    1996-03-01

    A novel laser-induced temperature jump (T-jump) of 20 C or more is used to initiate the unfolding process of peptides and proteins on the picosecond time scale, and amide I time-resolved infrared absorbance transients are used to characterize the resulting kinetics. We have used this method to study the kinetics of folding and unfolding of a small 21 residue alanine based peptide and molten globule and native states of apomyoglobin, models for the helix which is an basic motif found in proteins. An essential result of our study is that the folding kinetics of a short length of peptide can occur within a few tens of nanoseconds which is much shorter than the time scale of the formation of intramolecular tertiary contacts from one point of a polypeptide chain to another. Furthermore, we observed that helices stabilized by tertiary contact formation unfold slower than helices surrounded by solvent by three orders of magnitude. These results bear directly on the protein folding problem, that is how do proteins fold from a large number of heterogeneous unfolded states to find the specific biologically active folded state on biologically relevent time scales, by suggesting that secondary structure forms first followed by tertiary structure. This work is a collaborative effort with R. GILMANSHIN at City College and S. WILLIAMS, R. B. DYER, and W. H. WOODRUFF at CST-4, Los Alamos National Laboratory, Los Alamos, NM 87545.

  11. Dynamics of protein folding: probing the kinetic network of folding-unfolding transitions with experiment and theory.

    PubMed

    Buchner, Ginka S; Murphy, Ronan D; Buchete, Nicolae-Viorel; Kubelka, Jan

    2011-08-01

    The problem of spontaneous folding of amino acid chains into highly organized, biologically functional three-dimensional protein structures continues to challenge the modern science. Understanding how proteins fold requires characterization of the underlying energy landscapes as well as the dynamics of the polypeptide chains in all stages of the folding process. In recent years, important advances toward these goals have been achieved owing to the rapidly growing interdisciplinary interest and significant progress in both experimental techniques and theoretical methods. Improvements in the experimental time resolution led to determination of the timescales of the important elementary events in folding, such as formation of secondary structure and tertiary contacts. Sensitive single molecule methods made possible probing the distributions of the unfolded and folded states and following the folding reaction of individual protein molecules. Discovery of proteins that fold in microseconds opened the possibility of atomic-level theoretical simulations of folding and their direct comparisons with experimental data, as well as of direct experimental observation of the barrier-less folding transition. The ultra-fast folding also brought new questions, concerning the intrinsic limits of the folding rates and experimental signatures of barrier-less "downhill" folding. These problems will require novel approaches for even more detailed experimental investigations of the folding dynamics as well as for the analysis of the folding kinetic data. For theoretical simulations of folding, a main challenge is how to extract the relevant information from overwhelmingly detailed atomistic trajectories. New theoretical methods have been devised to allow a systematic approach towards a quantitative analysis of the kinetic network of folding-unfolding transitions between various configuration states of a protein, revealing the transition states and the associated folding pathways at

  12. Energy landscapes, folding mechanisms, and kinetics of RNA tetraloop hairpins.

    PubMed

    Chakraborty, Debayan; Collepardo-Guevara, Rosana; Wales, David J

    2014-12-31

    RNA hairpins play a pivotal role in a diverse range of cellular functions, and are integral components of ribozymes, mRNA, and riboswitches. However, the mechanistic and kinetic details of RNA hairpin folding, which are key determinants of most of its biological functions, are poorly understood. In this work, we use the discrete path sampling (DPS) approach to explore the energy landscapes of two RNA tetraloop hairpins, and provide insights into their folding mechanisms and kinetics in atomistic detail. Our results show that the potential energy landscapes have a distinct funnel-like bias toward the folded hairpin state, consistent with efficient structure-seeking properties. Mechanistic and kinetic information is analyzed in terms of kinetic transition networks. We find microsecond folding times, consistent with temperature jump experiments, for hairpin folding initiated from relatively compact unfolded states. This process is essentially driven by an initial collapse, followed by rapid zippering of the helix stem in the final phase. Much lower folding rates are predicted when the folding is initiated from extended chains, which undergo longer excursions on the energy landscape before nucleation events can occur. Our work therefore explains recent experiments and coarse-grained simulations, where the folding kinetics exhibit precisely this dependency on the initial conditions.

  13. Statistical mechanics of kinetic proofreading in protein folding in vivo.

    PubMed Central

    Gulukota, K; Wolynes, P G

    1994-01-01

    The statistical energy landscape picture of protein folding has led to the understanding that the energy landscape must have guiding forces leading to a protein folding funnel in order to avoid the Levinthal paradox in vitro. Since folding in vivo often requires the action of chaperone molecules and ATP hydrolysis, we must ask whether folding in a system maintained away from equilibrium can avoid the Levinthal paradox in other ways. We describe a model of the action of chaperone molecules in protein folding in vivo on the basis of a repetitive cycle of binding and unbinding, allowing the possibility of kinetic proofreading. We also study models in which chaperone binding is locally biased, depending on the similarity of the conformation to the native one. We show that while kinetic proofreading can modestly facilitate folding, it is insufficient by itself to overcome the Levinthal paradox. On the other hand, such kinetic proofreading with biasing can provide the nonequilibrium analog of a folding funnel and greatly enhance folding yields and speed up folding. PMID:7937758

  14. Length dependent folding kinetics of phenylacetylene oligomers: Structural characterization of a kinetic trap

    NASA Astrophysics Data System (ADS)

    Elmer, Sidney P.; Pande, Vijay S.

    2005-03-01

    Using simulation to study the folding kinetics of 20-mer poly-phenylacetylene (pPA) oligomers, we find a long time scale trapped kinetic phase in the cumulative folding time distribution. This is demonstrated using molecular dynamics to simulate an ensemble of over 100 folding trajectories. The simulation data are fit to a four-state kinetic model which includes the typical folded and unfolded states, along with an intermediate state, and most surprisingly, a kinetically trapped state. Topologically diverse conformations reminiscent of α helices, β turns, and sheets in proteins are observed, along with unique structures in the form of knots. The nonhelical conformations are implicated, on the basis of structural correlations to kinetic parameters, to contribute to the trapped kinetic behavior. The strong solvophobic forces which mediate the folding process and produce a stable helical folded state also serve to overstabilize the nonhelical conformations, ultimately trapping them. From our simulations, the folding time is predicted to be on the order of 2.5-12.5 μs in the presence of the trapped kinetic phase. The folding mechanism for these 20-mer chains is compared with the previously reported folding mechanism for the pPA 12-mer chains. A linear scaling relationship between the chain length and the mean first passage time is predicted in the absence of the trapped kinetic phase. We discuss the major implications of this discovery in the design of self-assembling nanostructures.

  15. Chemical, physical, and theoretical kinetics of an ultrafast folding protein.

    PubMed

    Kubelka, Jan; Henry, Eric R; Cellmer, Troy; Hofrichter, James; Eaton, William A

    2008-12-01

    An extensive set of equilibrium and kinetic data is presented and analyzed for an ultrafast folding protein--the villin subdomain. The equilibrium data consist of the excess heat capacity, tryptophan fluorescence quantum yield, and natural circular-dichroism spectrum as a function of temperature, and the kinetic data consist of time courses of the quantum yield from nanosecond-laser temperature-jump experiments. The data are well fit with three kinds of models--a three-state chemical-kinetics model, a physical-kinetics model, and an Ising-like theoretical model that considers 10(5) possible conformations (microstates). In both the physical-kinetics and theoretical models, folding is described as diffusion on a one-dimensional free-energy surface. In the physical-kinetics model the reaction coordinate is unspecified, whereas in the theoretical model, order parameters, either the fraction of native contacts or the number of native residues, are used as reaction coordinates. The validity of these two reaction coordinates is demonstrated from calculation of the splitting probability from the rate matrix of the master equation for all 10(5) microstates. The analysis of the data on site-directed mutants using the chemical-kinetics model provides information on the structure of the transition-state ensemble; the physical-kinetics model allows an estimate of the height of the free-energy barrier separating the folded and unfolded states; and the theoretical model provides a detailed picture of the free-energy surface and a residue-by-residue description of the evolution of the folded structure, yet contains many fewer adjustable parameters than either the chemical- or physical-kinetics models.

  16. Chemical, physical, and theoretical kinetics of an ultrafast folding protein

    PubMed Central

    Kubelka, Jan; Henry, Eric R.; Cellmer, Troy; Hofrichter, James; Eaton, William A.

    2008-01-01

    An extensive set of equilibrium and kinetic data is presented and analyzed for an ultrafast folding protein—the villin subdomain. The equilibrium data consist of the excess heat capacity, tryptophan fluorescence quantum yield, and natural circular-dichroism spectrum as a function of temperature, and the kinetic data consist of time courses of the quantum yield from nanosecond-laser temperature-jump experiments. The data are well fit with three kinds of models—a three-state chemical-kinetics model, a physical-kinetics model, and an Ising-like theoretical model that considers 105 possible conformations (microstates). In both the physical-kinetics and theoretical models, folding is described as diffusion on a one-dimensional free-energy surface. In the physical-kinetics model the reaction coordinate is unspecified, whereas in the theoretical model, order parameters, either the fraction of native contacts or the number of native residues, are used as reaction coordinates. The validity of these two reaction coordinates is demonstrated from calculation of the splitting probability from the rate matrix of the master equation for all 105 microstates. The analysis of the data on site-directed mutants using the chemical-kinetics model provides information on the structure of the transition-state ensemble; the physical-kinetics model allows an estimate of the height of the free-energy barrier separating the folded and unfolded states; and the theoretical model provides a detailed picture of the free-energy surface and a residue-by-residue description of the evolution of the folded structure, yet contains many fewer adjustable parameters than either the chemical- or physical-kinetics models. PMID:19033473

  17. Oxidative folding of peptides with cystine-knot architectures: kinetic studies and optimization of folding conditions.

    PubMed

    Reinwarth, Michael; Glotzbach, Bernhard; Tomaszowski, Michael; Fabritz, Sebastian; Avrutina, Olga; Kolmar, Harald

    2013-01-01

    Bioactive peptides often contain several disulfide bonds that provide the main contribution to conformational rigidity and structural, thermal, or biological stability. Among them, cystine-knot peptides-commonly named "knottins"-make up a subclass with several thousand natural members. Hence, they are considered promising frameworks for peptide-based pharmaceuticals. Although cystine-knot peptides are available through chemical and recombinant synthetic routes, oxidative folding to afford the bioactive isomers still remains a crucial step. We therefore investigated the oxidative folding of ten protease-inhibiting peptides from two knottin families, as well as that of an HIV entry inhibitor and of aprotinin, under two conventional sets of folding conditions and by a newly developed procedure. Kinetic studies identified folding conditions that resulted in correctly folded miniproteins with high rates of conversion even for highly hydrophobic and aggregation-prone peptides in concentrated solutions. PMID:23229141

  18. Periodic and stochastic thermal modulation of protein folding kinetics

    SciTech Connect

    Platkov, Max; Gruebele, Martin

    2014-07-21

    Chemical reactions are usually observed either by relaxation of a bulk sample after applying a sudden external perturbation, or by intrinsic fluctuations of a few molecules. Here we show that the two ideas can be combined to measure protein folding kinetics, either by periodic thermal modulation, or by creating artificial thermal noise that greatly exceeds natural thermal fluctuations. We study the folding reaction of the enzyme phosphoglycerate kinase driven by periodic temperature waveforms. As the temperature waveform unfolds and refolds the protein, its fluorescence color changes due to FRET (Förster resonant Energy Transfer) of two donor/acceptor fluorophores labeling the protein. We adapt a simple model of periodically driven kinetics that nicely fits the data at all temperatures and driving frequencies: The phase shifts of the periodic donor and acceptor fluorescence signals as a function of driving frequency reveal reaction rates. We also drive the reaction with stochastic temperature waveforms that produce thermal fluctuations much greater than natural fluctuations in the bulk. Such artificial thermal noise allows the recovery of weak underlying signals due to protein folding kinetics. This opens up the possibility for future detection of a stochastic resonance for protein folding subject to noise with controllable amplitude.

  19. Periodic and stochastic thermal modulation of protein folding kinetics

    NASA Astrophysics Data System (ADS)

    Platkov, Max; Gruebele, Martin

    2014-07-01

    Chemical reactions are usually observed either by relaxation of a bulk sample after applying a sudden external perturbation, or by intrinsic fluctuations of a few molecules. Here we show that the two ideas can be combined to measure protein folding kinetics, either by periodic thermal modulation, or by creating artificial thermal noise that greatly exceeds natural thermal fluctuations. We study the folding reaction of the enzyme phosphoglycerate kinase driven by periodic temperature waveforms. As the temperature waveform unfolds and refolds the protein, its fluorescence color changes due to FRET (Förster resonant Energy Transfer) of two donor/acceptor fluorophores labeling the protein. We adapt a simple model of periodically driven kinetics that nicely fits the data at all temperatures and driving frequencies: The phase shifts of the periodic donor and acceptor fluorescence signals as a function of driving frequency reveal reaction rates. We also drive the reaction with stochastic temperature waveforms that produce thermal fluctuations much greater than natural fluctuations in the bulk. Such artificial thermal noise allows the recovery of weak underlying signals due to protein folding kinetics. This opens up the possibility for future detection of a stochastic resonance for protein folding subject to noise with controllable amplitude.

  20. Microsecond Microfluidic Mixing for Investigation of Protein Folding Kinetics

    SciTech Connect

    Hertzog, D E; Santiago, J G; Bakajin, O

    2003-06-25

    We have developed and characterized a mixer to study the reaction kinetics of protein folding on a microsecond timescale. The mixer uses hydrodynamic focusing of pressure-driven flow in a microfluidic channel to reduce diffusion times as first demonstrated by Knight et al.[1]. Features of the mixer include 1 {micro}s mixing times, sample consumptions of order 1 nl/s, loading sample volumes on the order of microliters, and the ability to manufacture in fused silica for compatibility with most spectroscopic methods.

  1. Microsecond Microfluidic Mixing for Investigation of Protein Folding Kinetics

    SciTech Connect

    Hertzog, D E; Santiago, J G; Bakajin, O

    2005-02-10

    We have developed and characterized a mixer to study the reaction kinetics of protein folding on a microsecond timescale. The mixer uses hydrodynamic focusing of pressure-driven flow in a microfluidic channel to reduce diffusion times as first demonstrated by Knight et al.[1]. Features of the mixer include 1 {micro}s mixing times, sample consumptions of order 1 nl/s, loading sample volumes on the order of microliters, and the ability to manufacture in fused silica for compatibility with most spectroscopic methods.

  2. Combined approach to the inverse protein folding problem. Final report

    SciTech Connect

    Ruben A. Abagyan

    2000-06-01

    The main scientific contribution of the project ''Combined approach to the inverse protein folding problem'' submitted in 1996 and funded by the Department of Energy in 1997 is the formulation and development of the idea of the multilink recognition method for identification of functional and structural homologues of newly discovered genes. This idea became very popular after they first announced it and used it in prediction of the threading targets for the CASP2 competition (Critical Assessment of Structure Prediction).

  3. Configuration-dependent diffusion can shift the kinetic transition state and barrier height of protein folding

    PubMed Central

    Chahine, Jorge; Oliveira, Ronaldo J.; Leite, Vitor B. P.; Wang, Jin

    2007-01-01

    We show that diffusion can play an important role in protein-folding kinetics. We explicitly calculate the diffusion coefficient of protein folding in a lattice model. We found that diffusion typically is configuration- or reaction coordinate-dependent. The diffusion coefficient is found to be decreasing with respect to the progression of folding toward the native state, which is caused by the collapse to a compact state constraining the configurational space for exploration. The configuration- or position-dependent diffusion coefficient has a significant contribution to the kinetics in addition to the thermodynamic free-energy barrier. It effectively changes (increases in this case) the kinetic barrier height as well as the position of the corresponding transition state and therefore modifies the folding kinetic rates as well as the kinetic routes. The resulting folding time, by considering both kinetic diffusion and the thermodynamic folding free-energy profile, thus is slower than the estimation from the thermodynamic free-energy barrier with constant diffusion but is consistent with the results from kinetic simulations. The configuration- or coordinate-dependent diffusion is especially important with respect to fast folding, when there is a small or no free-energy barrier and kinetics is controlled by diffusion. Including the configurational dependence will challenge the transition state theory of protein folding. The classical transition state theory will have to be modified to be consistent. The more detailed folding mechanistic studies involving phi value analysis based on the classical transition state theory also will have to be modified quantitatively. PMID:17804812

  4. Translation Initiation is Controlled by RNA Folding Kinetics via a Ribosome Drafting Mechanism.

    PubMed

    Espah Borujeni, Amin; Salis, Howard M

    2016-06-01

    RNA folding plays an important role in controlling protein synthesis as well as other cellular processes. Existing models have focused on how RNA folding energetics control translation initiation rate under equilibrium conditions but have largely ignored the effects of nonequilibrium RNA folding. We introduce a new mechanism, called "ribosome drafting", that explains how a mRNA's folding kinetics and the ribosome's binding rate collectively control its translation initiation rate. During cycles of translation, ribosome drafting emerges whenever successive ribosomes bind to a mRNA faster than the mRNA can refold, maintaining it in a nonequilibrium state with an acceleration of protein synthesis. Using computational design, time-correlated single photon counting, and expression measurements, we demonstrate that slow-folding and fast-folding RNA structures with equivalent folding energetics can vary protein synthesis rates by 1000-fold. We determine the necessary conditions for ribosome drafting by characterizing mRNAs with rationally designed ribosome binding rates, folding kinetics, and folding energetics, confirming the predictions of a nonequilibrium Markov model of translation. Our results have widespread implications, illustrating how competitive folding and assembly kinetics can shape the gene expression machinery's sequence-structure-function relationship inside cells. PMID:27199273

  5. Solvent-Exposed Salt Bridges Influence the Kinetics of α-Helix Folding and Unfolding.

    PubMed

    Meuzelaar, Heleen; Tros, Martijn; Huerta-Viga, Adriana; van Dijk, Chris N; Vreede, Jocelyne; Woutersen, Sander

    2014-03-01

    Salt bridges are known to play an essential role in the thermodynamic stability of the folded conformation of many proteins, but their influence on the kinetics of folding remains largely unknown. Here, we investigate the effect of Glu-Arg salt bridges on the kinetics of α-helix folding using temperature-jump transient-infrared spectroscopy and steady-state UV circular dichroism. We find that geometrically optimized salt bridges (Glu(-) and Arg(+) are spaced four peptide units apart, and the Glu/Arg order is such that the side-chain rotameric preferences favor salt-bridge formation) significantly speed up folding and slow down unfolding, whereas salt bridges with unfavorable geometry slow down folding and slightly speed up unfolding. Our observations suggest a possible explanation for the surprising fact that many biologically active proteins contain salt bridges that do not stabilize the native conformation: these salt bridges might have a kinetic rather than a thermodynamic function.

  6. Monitoring the folding kinetics of a β-hairpin by time-resolved IR spectroscopy in silico.

    PubMed

    Daidone, Isabella; Thukral, Lipi; Smith, Jeremy C; Amadei, Andrea

    2015-04-01

    Protein folding is one of the most fundamental problems in modern biochemistry. Time-resolved infrared (IR) spectroscopy in the amide I region is commonly used to monitor folding kinetics. However, associated atomic detail information on the folding mechanism requires simulations. In atomistic simulations structural order parameters are typically used to follow the folding process along the simulated trajectories. However, a rigorous test of the reliability of the mechanisms found in the simulations requires calculation of the time-dependent experimental observable, i.e., in the present case the IR signal in the amide I region. Here, we combine molecular dynamics simulation with a mixed quantum mechanics/molecular mechanics theoretical methodology, the Perturbed Matrix Method, in order to characterize the folding of a β-hairpin peptide, through modeling the time-dependence of the amide I IR signal. The kinetic and thermodynamic data (folding and unfolding rate constants, and equilibrium folded- and unfolded-state probabilities) obtained from the fit of the calculated signal are in good agreement with the available experimental data [Xu et al. J. Am. Chem. Soc. 2003, 125, 15388-15394]. To the best of our knowledge, this is the first report of the simulation of the time-resolved IR signal of a complex process occurring on a long (microsecond) time scale.

  7. Monitoring the folding kinetics of a β-hairpin by time-resolved IR spectroscopy in silico.

    PubMed

    Daidone, Isabella; Thukral, Lipi; Smith, Jeremy C; Amadei, Andrea

    2015-04-01

    Protein folding is one of the most fundamental problems in modern biochemistry. Time-resolved infrared (IR) spectroscopy in the amide I region is commonly used to monitor folding kinetics. However, associated atomic detail information on the folding mechanism requires simulations. In atomistic simulations structural order parameters are typically used to follow the folding process along the simulated trajectories. However, a rigorous test of the reliability of the mechanisms found in the simulations requires calculation of the time-dependent experimental observable, i.e., in the present case the IR signal in the amide I region. Here, we combine molecular dynamics simulation with a mixed quantum mechanics/molecular mechanics theoretical methodology, the Perturbed Matrix Method, in order to characterize the folding of a β-hairpin peptide, through modeling the time-dependence of the amide I IR signal. The kinetic and thermodynamic data (folding and unfolding rate constants, and equilibrium folded- and unfolded-state probabilities) obtained from the fit of the calculated signal are in good agreement with the available experimental data [Xu et al. J. Am. Chem. Soc. 2003, 125, 15388-15394]. To the best of our knowledge, this is the first report of the simulation of the time-resolved IR signal of a complex process occurring on a long (microsecond) time scale. PMID:25777154

  8. Effects of mutation, truncation, and temperature on the folding kinetics of a WW domain.

    PubMed

    Maisuradze, Gia G; Zhou, Rui; Liwo, Adam; Xiao, Yi; Scheraga, Harold A

    2012-07-20

    The purpose of this work is to show how mutation, truncation, and change of temperature can influence the folding kinetics of a protein. This is accomplished by principal component analysis of molecular-dynamics-generated folding trajectories of the triple β-strand WW domain from formin binding protein 28 (FBP28) (Protein Data Bank ID: 1E0L) and its full-size, and singly- and doubly-truncated mutants at temperatures below and very close to the melting point. The reasons for biphasic folding kinetics [i.e., coexistence of slow (three-state) and fast (two-state) phases], including the involvement of a solvent-exposed hydrophobic cluster and another delocalized hydrophobic core in the folding kinetics, are discussed. New folding pathways are identified in free-energy landscapes determined in terms of principal components for full-size mutants. Three-state folding is found to be a main mechanism for folding the FBP28 WW domain and most of the full-size and truncated mutants. The results from the theoretical analysis are compared to those from experiment. Agreements and discrepancies between the theoretical and experimental results are discussed. Because of its importance in understanding protein kinetics and function, the diffusive mechanism by which the FBP28 WW domain and its full-size and truncated mutants explore their conformational space is examined in terms of the mean-square displacement and principal component analysis eigenvalue spectrum analyses. Subdiffusive behavior is observed for all studied systems. PMID:22560992

  9. Microscopic dynamics from a coarsely defined solution to the protein folding problem

    NASA Astrophysics Data System (ADS)

    Fernández, Ariel; Colubri, Andrés

    1998-06-01

    local torsional constraints. The validity of our approach is tested vis à vis experimentally probed folding pathways generating tertiary interactions in proteins that may recover their active structure under in vitro renaturation conditions. We focus on determining significant folding intermediates and the late kinetic bottlenecks that occur within the first 1/100 s of the renaturation process. After the computational accessibility of this coarse solution of the folding problem becomes apparent, we show how to lift our variational problem to microscopic dynamics of the peptide chain. The consistency of our approach is revealed by an actual generation of Newton's laws at the microscopic level through an inverse projection of the coarse dynamics originally generated through the pattern recognition computation.

  10. Kinetics methods for clinical epidemiology problems

    PubMed Central

    Corlan, Alexandru Dan; Ross, John

    2015-01-01

    Calculating the probability of each possible outcome for a patient at any time in the future is currently possible only in the simplest cases: short-term prediction in acute diseases of otherwise healthy persons. This problem is to some extent analogous to predicting the concentrations of species in a reactor when knowing initial concentrations and after examining reaction rates at the individual molecule level. The existing theoretical framework behind predicting contagion and the immediate outcome of acute diseases in previously healthy individuals is largely analogous to deterministic kinetics of chemical systems consisting of one or a few reactions. We show that current statistical models commonly used in chronic disease epidemiology correspond to simple stochastic treatment of single reaction systems. The general problem corresponds to stochastic kinetics of complex reaction systems. We attempt to formulate epidemiologic problems related to chronic diseases in chemical kinetics terms. We review methods that may be adapted for use in epidemiology. We show that some reactions cannot fit into the mass-action law paradigm and solutions to these systems would frequently exhibit an antiportfolio effect. We provide a complete example application of stochastic kinetics modeling for a deductive meta-analysis of two papers on atrial fibrillation incidence, prevalence, and mortality. PMID:26578757

  11. Kinetics methods for clinical epidemiology problems.

    PubMed

    Corlan, Alexandru Dan; Ross, John

    2015-11-17

    Calculating the probability of each possible outcome for a patient at any time in the future is currently possible only in the simplest cases: short-term prediction in acute diseases of otherwise healthy persons. This problem is to some extent analogous to predicting the concentrations of species in a reactor when knowing initial concentrations and after examining reaction rates at the individual molecule level. The existing theoretical framework behind predicting contagion and the immediate outcome of acute diseases in previously healthy individuals is largely analogous to deterministic kinetics of chemical systems consisting of one or a few reactions. We show that current statistical models commonly used in chronic disease epidemiology correspond to simple stochastic treatment of single reaction systems. The general problem corresponds to stochastic kinetics of complex reaction systems. We attempt to formulate epidemiologic problems related to chronic diseases in chemical kinetics terms. We review methods that may be adapted for use in epidemiology. We show that some reactions cannot fit into the mass-action law paradigm and solutions to these systems would frequently exhibit an antiportfolio effect. We provide a complete example application of stochastic kinetics modeling for a deductive meta-analysis of two papers on atrial fibrillation incidence, prevalence, and mortality.

  12. Kinetically coupled folding of a single HIV-1 glycoprotein 41 complex in viral membrane fusion and inhibition

    PubMed Central

    Jiao, Junyi; Rebane, Aleksander A.; Ma, Lu; Gao, Ying; Zhang, Yongli

    2015-01-01

    HIV-1 glycoprotein 41 (gp41) mediates viral entry into host cells by coupling its folding energy to membrane fusion. Gp41 folding is blocked by fusion inhibitors, including the commercial drug T20, to treat HIV/AIDS. However, gp41 folding intermediates, energy, and kinetics are poorly understood. Here, we identified the folding intermediates of a single gp41 trimer-of-hairpins and measured their associated energy and kinetics using high-resolution optical tweezers. We found that folding of gp41 hairpins was energetically independent but kinetically coupled: Each hairpin contributed a folding energy of ∼−23 kBT, but folding of one hairpin successively accelerated the folding rate of the next one by ∼20-fold. Membrane-mimicking micelles slowed down gp41 folding and reduced the stability of the six-helix bundle. However, the stability was restored by cooperative folding of the membrane-proximal external region. Surprisingly, T20 strongly inhibited gp41 folding by actively displacing the C-terminal hairpin strand in a force-dependent manner. The inhibition was abolished by a T20-resistant gp41 mutation. The energetics and kinetics of gp41 folding established by us provides a basis to understand viral membrane fusion, infection, and therapeutic intervention. PMID:26038562

  13. Deconstructing time-resolved optical rotatory dispersion kinetic measurements of cytochrome c folding: from molten globule to the native state.

    PubMed

    Chen, Eefei; Kliger, David S

    2012-01-01

    The far-UV time-resolved optical rotatory dispersion (TRORD) technique has contributed significantly to our understanding of nanosecond secondary structure kinetics in protein folding and function reactions. For reduced cytochrome c, protein folding kinetics have been probed largely from the unfolded to the native state. Here we provide details about sample preparation and the TRORD apparatus and measurements for studying folding from a partly unfolded state to the native secondary structure conformation of reduced cytochrome c. PMID:22760330

  14. The initial step of DNA hairpin folding: a kinetic analysis using fluorescence correlation spectroscopy

    PubMed Central

    Kim, Jiho; Doose, Sören; Neuweiler, Hannes; Sauer, Markus

    2006-01-01

    Conformational fluctuations of single-stranded DNA (ssDNA) oligonucleotides were studied in aqueous solution by monitoring contact-induced fluorescence quenching of the oxazine fluorophore MR121 by intrinsic guanosine residues (dG). We applied fluorescence correlation spectroscopy as well as steady-state and time-resolved fluorescence spectroscopy to analyze kinetics of DNA hairpin folding. We first characterized the reporter system by investigating bimolecular quenching interactions between MR121 and guanosine monophosphate in aqueous solution estimating rate constants, efficiency and stability for formation of quenched complexes. We then studied the kinetics of complex formation between MR121 and dG residues site-specifically incorporated in DNA hairpins. To uncover the initial steps of DNA hairpin folding we investigated complex formation in ssDNA carrying one or two complementary base pairs (dC–dG pairs) that could hybridize to form a short stem. Our data show that incorporation of a single dC–dG pair leads to non-exponential decays for opening and closing kinetics and reduces rate constants by one to two orders of magnitude. We found positive activation enthalpies independent of the number of dC–dG pairs. These results imply that the rate limiting step of DNA hairpin folding is not determined by loop dynamics, or by mismatches in the stem, but rather by interactions between stem and loop nucleotides. PMID:16687657

  15. ETNA: equilibrium transitions network and Arrhenius equation for extracting folding kinetics from REMD simulations.

    PubMed

    Muff, S; Caflisch, A

    2009-03-12

    It is difficult to investigate folding kinetics by conventional atomistic simulations of proteins. The replica exchange molecular dynamics (REMD) simulation technique enhances conformational sampling at the expenses of reduced kinetic information, which in REMD is directly available only for very short time scales. Here, we propose a procedure for obtaining kinetic data from REMD by making use of the equilibrium transitions network (ETN) sampled at the temperature of interest. This information is supplemented by mean folding times extracted from ETNs at higher REMD temperatures and scaled according to the Arrhenius equation. The procedure is applied to a three-stranded antiparallel beta-sheet peptide which has a very heterogeneous denatured state with a broad entropic basin and several enthalpic traps. Despite the complexity of the system and the REMD exchange time of only 0.1 ns, the procedure is able to estimate folding times (ranging from about 0.1 micros at the melting temperature of 330 K to about 8 micros at 286 K) as well as transition times from individual non-native basins to the native state.

  16. A combined kinetic push and thermodynamic pull as driving forces for outer membrane protein sorting and folding in bacteria

    PubMed Central

    Fleming, Karen G.

    2015-01-01

    In vitro folding studies of outer membrane beta-barrels have been invaluable in revealing the lipid effects on folding rates and efficiencies as well as folding free energies. Here, the biophysical results are summarized, and these kinetic and thermodynamic findings are considered in terms of the requirements for folding in the context of the cellular environment. Because the periplasm lacks an external energy source the only driving forces for sorting and folding available within this compartment are binding or folding free energies and their associated rates. These values define functions for periplasmic chaperones and suggest a biophysical mechanism for the BAM complex. PMID:26370938

  17. Kinetic barriers to the folding of horse cytochrome C in the reduced state.

    PubMed

    Bhuyan, Abani K; Kumar, Rajesh

    2002-10-22

    To determine the kinetic barrier in the folding of horse cytochrome c, a CO-liganded derivative of cytochrome c, called carbonmonoxycytochrome c, has been prepared by exploiting the thermodynamic reversibility of ferrocytochrome c unfolding induced by guanidinium hydrochloride (GdnHCl), pH 7. The CO binding properties of unfolded ferrocytochrome c, studied by 13C NMR and optical spectroscopy, are remarkably similar to those of native myoglobin and isolated chains of human hemoglobin. Equilibrium unfolding transitions of ferrocytochrome c in the presence and the absence of CO observed by both excitation energy transfer from the lone tryptophan to the ferrous heme and far-UV circular dichroism (CD) indicate no accumulation of structural intermediates to a detectable level. Values of thermodynamic parameters obtained by two-state analysis of fluorescence transitions are DeltaG(H2O) = 11.65(+/-1.13) kcal x mol(-1) and C(m) = 3.9(+/-0.1) M GdnHCl in the presence of CO, and DeltaG(H2O)=19.3(+/-0.5) kcal x mol(-1) and C(m) = 5.1(+/-0.1) M GdnHCl in the absence of CO, indicating destabilization of ferrocytochrome c by approximately 7.65 kcal x mol(-1) due to CO binding. The native states of ferrocytochrome c and carbonmonoxycytochrome c are nearly identical in terms of structure and conformation except for the Fe2+-M80 --> Fe2+-CO replacement. Folding and unfolding kinetics as a function of GdnHCl, studied by stopped-flow fluorescence, are significantly different for the two proteins. Both refold fast, but carbonmonoxycytochrome c refolds 2-fold faster (tau = 1092 micros at 10 degrees C) than ferrocytochrome c. Linear extrapolation of the folding rates to the ordinate of the chevron plot projects this value of tau to 407 micros. The unfolding rate of the former in water, estimated by extrapolation, is faster by more than 10 orders of magnitude. Significant differences are also observed in rate-denaturant gradients in the chevron. Formation and disruption of the Fe2+-M80

  18. Estimating free-energy barrier heights for an ultrafast folding protein from calorimetric and kinetic data.

    PubMed

    Godoy-Ruiz, Raquel; Henry, Eric R; Kubelka, Jan; Hofrichter, James; Muñoz, Victor; Sanchez-Ruiz, Jose M; Eaton, William A

    2008-05-15

    Differential scanning calorimetry was used to measure the temperature dependence of the absolute heat capacity of the 35-residue subdomain of the villin headpiece, a protein that folds in 5 mus and is therefore assumed to have a small free-energy barrier separating folded and unfolded states. To obtain an estimate of the barrier height from the calorimetric data, two models, a variable-barrier model and an Ising-like model, were used to fit the heat capacity in excess of the folded state over the temperature range 15-125 degrees C. The variable-barrier model is based on an empirical mathematical form for the density of states, with four adjustable parameters and the enthalpy (H) as a reaction coordinate. The Ising-like model is based on the inter-residue contact map of the X-ray structure with exact enumeration of approximately 10(5) possible conformations, with two adjustable parameters in the partition function, and either the fraction of native contacts (Q) or the number of ordered residues (P) as reaction coordinates. The variable-barrier model provides an excellent fit to the data and yields a barrier height at the folding temperature ranging from 0.4 to 1.1 kcal mol(-1), while the Ising-like model provides a less good fit and yields barrier heights of 2.3 +/- 0.1 kcal mol(-1) and 2.1 +/- 0.1 kcal mol(-1) for the Q and P reaction coordinates, respectively. In both models, the barrier to folding increases with increasing temperature. Assuming a sufficiently large activation energy for diffusion on the free-energy surfaces, both models are consistent with the observation of a temperature-independent folding rate in previously published laser temperature-jump experiments. Analysis of this kinetic data, using an approximate form for the pre-exponential factor of Kramers theory and the 70 ns relaxation time for the fast phase that precedes the unfolding/refolding relaxation to determine the diffusion coefficient, results in a barrier height of 1.6 +/- 0.3 kcal mol

  19. Computational design of RNA parts, devices, and transcripts with kinetic folding algorithms implemented on multiprocessor clusters.

    PubMed

    Thimmaiah, Tim; Voje, William E; Carothers, James M

    2015-01-01

    With progress toward inexpensive, large-scale DNA assembly, the demand for simulation tools that allow the rapid construction of synthetic biological devices with predictable behaviors continues to increase. By combining engineered transcript components, such as ribosome binding sites, transcriptional terminators, ligand-binding aptamers, catalytic ribozymes, and aptamer-controlled ribozymes (aptazymes), gene expression in bacteria can be fine-tuned, with many corollaries and applications in yeast and mammalian cells. The successful design of genetic constructs that implement these kinds of RNA-based control mechanisms requires modeling and analyzing kinetically determined co-transcriptional folding pathways. Transcript design methods using stochastic kinetic folding simulations to search spacer sequence libraries for motifs enabling the assembly of RNA component parts into static ribozyme- and dynamic aptazyme-regulated expression devices with quantitatively predictable functions (rREDs and aREDs, respectively) have been described (Carothers et al., Science 334:1716-1719, 2011). Here, we provide a detailed practical procedure for computational transcript design by illustrating a high throughput, multiprocessor approach for evaluating spacer sequences and generating functional rREDs. This chapter is written as a tutorial, complete with pseudo-code and step-by-step instructions for setting up a computational cluster with an Amazon, Inc. web server and performing the large numbers of kinefold-based stochastic kinetic co-transcriptional folding simulations needed to design functional rREDs and aREDs. The method described here should be broadly applicable for designing and analyzing a variety of synthetic RNA parts, devices and transcripts.

  20. Quantitative analysis of the kinetics of denaturation and renaturation of barstar in the folding transition zone.

    PubMed Central

    Shastry, M. C.; Agashe, V. R.; Udgaonkar, J. B.

    1994-01-01

    The fluorescence-monitored kinetics of folding and unfolding of barstar by guanidine hydrochloride (GdnHCl) in the folding transition zone, at pH 7, 25 degrees C, have been quantitatively analyzed using a 3-state mechanism: U(S)<-->UF<-->N. U(S) and UF are slow-refolding and fast-refolding unfolded forms of barstar, and N is the native protein. U(S) and UF probably differ in possessing trans and cis conformations, respectively, of the Tyr 47-Pro 48 bond. The 3-state model could be used because the kinetics of folding and unfolding of barstar show 2 phases, a fast phase and a slow phase, and because the relative amplitudes of the 2 phases depend only on the final refolding conditions and not on the initial conditions. Analysis of the observed kinetics according to the 3-state model yields the values of the 4 microscopic rate constants that describe the transitions between the 3 states at different concentrations of GdnHCl. The value of the equilibrium unfolded ratio U(S):UF (K21) and the values of the rate constants of the U(S)-->UF and UF-->U(S) reactions, k12 and k21, respectively, are shown to be independent of the concentration of GdnHCl. K21 has a value of 2.1 +/- 0.1, and k12 and k21 have values of 5.3 x 10(-3) s-1 and 11.2 x 10(-3) s-1, respectively. Double-jump experiments that monitor reactions that are silent to fluorescence monitoring were used to confirm the values of K21, k12, and k21 obtained from the 3-state analysis and thereby the validity of the 3-state model.(ABSTRACT TRUNCATED AT 250 WORDS) PMID:7833803

  1. Single-molecule kinetics under force: probing protein folding and enzymatic activity with optical tweezers

    NASA Astrophysics Data System (ADS)

    Wong, Wesley

    2010-03-01

    Weak non-covalent bonds between and within single molecules govern many aspects of biological structure and function (e.g. DNA base-paring, receptor-ligand binding, protein folding, etc.) In living systems, these interactions are often subject to mechanical forces, which can greatly alter their kinetics and activity. My group develops and applies novel single-molecule manipulation techniques to explore and quantify these force-dependent kinetics. Using optical tweezers, we have quantified the force-dependent unfolding and refolding kinetics of different proteins, including the cytoskeletal protein spectrin in collaboration with E. Evans's group [1], and the A2 domain of the von Willebrand factor blood clotting protein in collaboration with T. Springer's group [2]. Furthermore, we have studied the kinetics of the ADAMTS13 enzyme acting on a single A2 domain, and have shown that physiolgical forces in the circulation can act as a cofactor for enzymatic cleavage, regulating hemostatic activity [2]. References: 1. E. Evans, K. Halvorsen, K. Kinoshita, and W.P. Wong, Handbook of Single Molecule Biophysics, P. Hinterdorfer, ed., Springer (2009). 2. X. Zhang, K. Halvorsen, C.-Z. Zhang, W.P. Wong, and T.A. Springer, Science 324 (5932), 1330-1334 (2009).

  2. Local Kinetic Measures of Macromolecular Structure Reveal Partitioning Among Multiple Parallel Pathways from the Earliest Steps in the Folding of a Large RNA Molecule

    SciTech Connect

    Laederach,A.; Shcherbakova, I.; Liang, M.; Brenowitz, M.; Altman, R.

    2006-01-01

    At the heart of the RNA folding problem is the number, structures, and relationships among the intermediates that populate the folding pathways of most large RNA molecules. Unique insight into the structural dynamics of these intermediates can be gleaned from the time-dependent changes in local probes of macromolecular conformation (e.g. reports on individual nucleotide solvent accessibility offered by hydroxyl radical ({center_dot}OH) footprinting). Local measures distributed around a macromolecule individually illuminate the ensemble of separate changes that constitute a folding reaction. Folding pathway reconstruction from a multitude of these individual measures is daunting due to the combinatorial explosion of possible kinetic models as the number of independent local measures increases. Fortunately, clustering of time progress curves sufficiently reduces the dimensionality of the data so as to make reconstruction computationally tractable. The most likely folding topology and intermediates can then be identified by exhaustively enumerating all possible kinetic models on a super-computer grid. The folding pathways and measures of the relative flux through them were determined for Mg{sup 2+} and Na{sup +}-mediated folding of the Tetrahymena thermophila group I intron using this combined experimental and computational approach. The flux during Mg{sup 2+}-mediated folding is divided among numerous parallel pathways. In contrast, the flux during the Na{sup +}-mediated reaction is predominantly restricted through three pathways, one of which is without detectable passage through intermediates. Under both conditions, the folding reaction is highly parallel with no single pathway accounting for more than 50% of the molecular flux. This suggests that RNA folding is non-sequential under a variety of different experimental conditions even at the earliest stages of folding. This study provides a template for the systematic analysis of the time-evolution of RNA structure

  3. Kinetic Dissection of the Pre-existing Conformational Equilibrium in the Trypsin Fold*

    PubMed Central

    Vogt, Austin D.; Chakraborty, Pradipta; Di Cera, Enrico

    2015-01-01

    Structural biology has recently documented the conformational plasticity of the trypsin fold for both the protease and zymogen in terms of a pre-existing equilibrium between closed (E*) and open (E) forms of the active site region. How such plasticity is manifested in solution and affects ligand recognition by the protease and zymogen is poorly understood in quantitative terms. Here we dissect the E*-E equilibrium with stopped-flow kinetics in the presence of excess ligand or macromolecule. Using the clotting protease thrombin and its zymogen precursor prethrombin-2 as relevant models we resolve the relative distribution of the E* and E forms and the underlying kinetic rates for their interconversion. In the case of thrombin, the E* and E forms are distributed in a 1:4 ratio and interconvert on a time scale of 45 ms. In the case of prethrombin-2, the equilibrium is shifted strongly (10:1 ratio) in favor of the closed E* form and unfolds over a faster time scale of 4.5 ms. The distribution of E* and E forms observed for thrombin and prethrombin-2 indicates that zymogen activation is linked to a significant shift in the pre-existing equilibrium between closed and open conformations that facilitates ligand binding to the active site. These findings broaden our mechanistic understanding of how conformational transitions control ligand recognition by thrombin and its zymogen precursor prethrombin-2 and have direct relevance to other members of the trypsin fold. PMID:26216877

  4. Integral and differential form of the protein folding problem

    NASA Astrophysics Data System (ADS)

    Tramontano, Anna

    2004-07-01

    The availability of the complete genomic sequences of many species, including human, has raised enormous expectations in medicine, pharmacology, ecology, biotechnology and forensic sciences. However, knowledge is only a first step toward understanding, and we are only at the early stage of a scientific process that might lead us to satisfy all the expectations raised by the genomic projects. In this review I will discuss the present status of computational methods that attempt to infer the unique three-dimensional structure of proteins from their amino acid sequences. Although this problem has been defined as the “holy grail” of biology, it represents only one of the many hurdles in our path towards the understanding of life at a molecular level.

  5. Kinetic Definition of Protein Folding Transition State Ensembles and Reaction Coordinates

    PubMed Central

    Snow, Christopher D.; Rhee, Young Min; Pande, Vijay S.

    2006-01-01

    Using distributed molecular dynamics simulations we located four distinct folding transitions for a 39-residue ββαβ protein fold. To characterize the nature of each room temperature transition, we calculated the probability of transmission for 500 points along each free energy barrier. We introduced a method for determining transition states by employing the transmission probability, Ptrans, and determined which conformations were transition state ensemble members (Ptrans ≈ 0.5). The transmission probability may be used to characterize the barrier in several ways. For example, we ran simulations at 82°C, determined the change in Ptrans with temperature for all 2,000 conformations, and quantified Hammond behavior directly using Ptrans correlation. Additionally, we propose that diffusion along Ptrans may provide the configurational diffusion rate at the top of the barrier. Specifically, given a transition state conformation x0 with estimated Ptrans = 0.5, we selected a large set of subsequent conformations from independent trajectories, each exactly a small time δt after x0 (250 ps). Calculating Ptrans for the new trial conformations, we generated the P(Ptrans|δt = 250 ps) distribution that reflected diffusion. This approach provides a novel perspective on the diffusive nature of a protein folding transition and provides a framework for a quantitative study of activated relaxation kinetics. PMID:16617068

  6. Concordant Exploration of the Kinetics of RNA Folding from Global and Local Perspectives

    SciTech Connect

    Kwok,L.; Scherbakova, I.; Lamb, J.; Park, H.; Andresen, K.; Smith, H.; Brenowitz, M.; Pollack, L.

    2006-01-01

    Time-resolved small-angle X-ray scattering (SAXS) with millisecond time-resolution reveals two discrete phases of global compaction upon Mg{sup 2+}-mediated folding of the Tetrahymena thermophila ribozyme. Electrostatic relaxation of the RNA occurs rapidly and dominates the first phase of compaction during which the observed radius of gyration (R{sub g}) decreases from 75 Angstroms to 55 Angstroms. A further decrease in R{sub g} to 45 Angstroms occurs in a well-defined second phase. An analysis of mutant ribozymes shows that the latter phase depends upon the formation of long-range tertiary contacts within the P4-P6 domain of the ribozyme; disruption of the three remaining long-range contacts linking the peripheral helices has no effect on the 55-45 Angstroms compaction transition. A better understanding of the role of specific tertiary contacts in compaction was obtained by concordant time-resolved hydroxyl radical ({center_dot}OH) analyses that report local changes in the solvent accessibility of the RNA backbone. Comparison of the global and local measures of folding shows that formation of a subset of native tertiary contacts (i.e. those defining the ribozyme core) can occur within a highly compact ensemble whose R{sub g} is close to that of the fully folded ribozyme. Analyses of additional ribozyme mutants and reaction conditions establish the generality of the rapid formation of a partially collapsed state with little to no detectable tertiary structure. These studies directly link global RNA compaction with formation of tertiary structure as the molecule acquires its biologically active structure, and underscore the strong dependence on salt of both local and global measures of folding kinetics.

  7. Improvements in Mixing Time and Mixing Uniformity in Devices Designed for Studies of Protein Folding Kinetics

    SciTech Connect

    Yao, Shuhuai; Bakajin, Olgica

    2007-08-01

    Using a microfluidic laminar flow mixer designed for studies of protein folding kinetics, we demonstrate a mixing time of 1 +/- 1 micros with sample consumption on the order of femtomoles. We recognize two limitations of previously proposed designs: (1) size and shape of the mixing region, which limits mixing uniformity and (2) the formation of Dean vortices at high flow rates, which limits the mixing time. We address these limitations by using a narrow shape-optimized nozzle and by reducing the bend of the side channel streamlines. The final design, which combines both of these features, achieves the best performance. We quantified the mixing performance of the different designs by numerical simulation of coupled Navier-Stokes and convection-diffusion equations and experiments using fluorescence resonance energy-transfer (FRET)-labeled DNA.

  8. Entanglement in correlated random spin chains, RNA folding and kinetic roughening

    NASA Astrophysics Data System (ADS)

    Rodríguez-Laguna, Javier; Santalla, Silvia N.; Ramírez, Giovanni; Sierra, Germán

    2016-07-01

    Average block entanglement in the 1D XX-model with uncorrelated random couplings is known to grow as the logarithm of the block size, in similarity to conformal systems. In this work we study random spin chains whose couplings present long range correlations, generated as gaussian fields with a power-law spectral function. Ground states are always planar valence bond states, and their statistical ensembles are characterized in terms of their block entropy and their bond-length distribution, which follow power-laws. We conjecture the existence of a critical value for the spectral exponent, below which the system behavior is identical to the case of uncorrelated couplings. Above that critical value, the entanglement entropy violates the area law and grows as a power law of the block size, with an exponent which increases from zero to one. Interestingly, we show that XXZ models with positive anisotropy present the opposite behavior, and strong correlations in the couplings lead to lower entropies. Similar planar bond structures are also found in statistical models of RNA folding and kinetic roughening, and we trace an analogy between them and quantum valence bond states. Using an inverse renormalization procedure we determine the optimal spin-chain couplings which give rise to a given planar bond structure, and study the statistical properties of the couplings whose bond structures mimic those found in RNA folding.

  9. Folding kinetics of WW domains with the united residue force field for bridging microscopic motions and experimental measurements

    PubMed Central

    Zhou, Rui; Maisuradze, Gia G.; Suñol, David; Todorovski, Toni; Macias, Maria J.; Xiao, Yi; Scheraga, Harold A.; Czaplewski, Cezary; Liwo, Adam

    2014-01-01

    To demonstrate the utility of the coarse-grained united-residue (UNRES) force field to compare experimental and computed kinetic data for folding proteins, we have performed long-time millisecond-timescale canonical Langevin molecular dynamics simulations of the triple β-strand from the Formin binding protein 28 WW domain and six nonnatural variants, using UNRES. The results have been compared with available experimental data in both a qualitative and a quantitative manner. Complexities of the folding pathways, which cannot be determined experimentally, were revealed. The folding mechanisms obtained from the simulated folding kinetics are in agreement with experimental results, with a few discrepancies for which we have accounted. The origins of single- and double-exponential kinetics and their correlations with two- and three-state folding scenarios are shown to be related to the relative barrier heights between the various states. The rate constants obtained from time profiles of the fractions of the native, intermediate, and unfolded structures, and the kinetic equations fitted to them, correlate with the experimental values; however, they are about three orders of magnitude larger than the experimental ones for most of the systems. These differences are in agreement with the timescale extension derived by scaling down the friction of water and averaging out the fast degrees of freedom when passing from all-atom to a coarse-grained representation. Our results indicate that the UNRES force field can provide accurate predictions of folding kinetics of these WW domains, often used as models for the study of the mechanisms of proein folding. PMID:25489078

  10. Redesigning the type II' β-turn in green fluorescent protein to type I': implications for folding kinetics and stability.

    PubMed

    Madan, Bharat; Sokalingam, Sriram; Raghunathan, Govindan; Lee, Sun-Gu

    2014-10-01

    Both Type I' and Type II' β-turns have the same sense of the β-turn twist that is compatible with the β-sheet twist. They occur predominantly in two residue β-hairpins, but the occurrence of Type I' β-turns is two times higher than Type II' β-turns. This suggests that Type I' β-turns may be more stable than Type II' β-turns, and Type I' β-turn sequence and structure can be more favorable for protein folding than Type II' β-turns. Here, we redesigned the native Type II' β-turn in GFP to Type I' β-turn, and investigated its effect on protein folding and stability. The Type I' β-turns were designed based on the statistical analysis of residues in natural Type I' β-turns. The substitution of the native "GD" sequence of i+1 and i+2 residues with Type I' preferred "(N/D)G" sequence motif increased the folding rate by 50% and slightly improved the thermodynamic stability. Despite the enhancement of in vitro refolding kinetics and stability of the redesigned mutants, they showed poor soluble expression level compared to wild type. To overcome this problem, i and i + 3 residues of the designed Type I' β-turn were further engineered. The mutation of Thr to Lys at i + 3 could restore the in vivo soluble expression of the Type I' mutant. This study indicates that Type II' β-turns in natural β-hairpins can be further optimized by converting the sequence to Type I'.

  11. Folding mechanism of reduced cytochrome c: Equilibrium and kinetic properties in the presence of carbon monoxide

    PubMed Central

    Latypov, Ramil F.; Maki, Kosuke; Cheng, Hong; Luck, Stanley D.; Roder, Heinrich

    2008-01-01

    Despite close structural similarity, the ferric and ferrous forms of cytochrome c (cyt c) differ greatly in terms of their ligand binding properties, stability, folding and dynamics. The reduced heme iron binds diatomic ligands such as CO only under destabilizing conditions that promote weakening or disruption of the native methionine-iron linkage. This makes CO a useful conformational probe for detecting partially structured states that cannot be observed in the absence of endogenous ligands. Heme absorbance, circular dichroism and NMR were used to characterize the denaturant-induced unfolding equilibrium of Fe2+ cyt c in the presence and absence of CO. In addition to the native state (N), which does not bind CO, and the unfolded CO-complex (U-CO), a structurally distinct CO-bound form (M-CO) accumulates to high levels (~75% of the population) at intermediate guanidine hydrochloride concentrations. Comparison of the unfolding transition for different conformational probes reveals that M-CO is a compact state containing a native-like helical core and regions of local disorder in the segment containing the native Met80 ligand and adjacent loops. Kinetic measurements of CO binding and dissociation under native, partially denaturing and fully unfolded conditions indicate that a state, M, that is structurally analogous to M-CO is populated even in the absence of CO. The binding energy of the CO ligand lowers the free energy of this high-energy state to such an extent that it accumulates even under mildly denaturing equilibrium conditions. The thermodynamic and kinetic parameters obtained in this study provide a fully self-consistent description of the linked unfolding/CO-binding equilibria of reduced cyt c. PMID:18761351

  12. Approximate Solutions for a Self-Folding Problem of Carbon Nanotubes

    SciTech Connect

    Y Mikata

    2006-08-22

    This paper treats approximate solutions for a self-folding problem of carbon nanotubes. It has been observed in the molecular dynamics calculations [1] that a carbon nanotube with a large aspect ratio can self-fold due to van der Waals force between the parts of the same carbon nanotube. The main issue in the self-folding problem is to determine the minimum threshold length of the carbon nanotube at which it becomes possible for the carbon nanotube to self-fold due to the van der Waals force. An approximate mathematical model based on the force method is constructed for the self-folding problem of carbon nanotubes, and it is solved exactly as an elastica problem using elliptic functions. Additionally, three other mathematical models are constructed based on the energy method. As a particular example, the lower and upper estimates for the critical threshold (minimum) length are determined based on both methods for the (5,5) armchair carbon nanotube.

  13. Trigger factor slows co-translational folding through kinetic trapping while sterically protecting the nascent chain from aberrant cytosolic interactions.

    PubMed

    O'Brien, Edward P; Christodoulou, John; Vendruscolo, Michele; Dobson, Christopher M

    2012-07-01

    The E. coli chaperone trigger factor (TF) interacts directly with nascent polypeptide chains as they emerge from the ribosome exit tunnel. Small protein domains can fold under the cradle created by TF, but the co-translational folding of larger proteins is slowed down by its presence. Because of the great experimental challenges in achieving high spatial and time resolution, it is not yet known whether or not TF alters the folding properties of small proteins and if the reduced rate of folding of larger proteins is the result of kinetic or thermodynamic effects. We show, by molecular simulations employing a coarse-grained model of a series of ribosome nascent-chain complexes, that TF does not alter significantly the co-translational folding process of a small protein G domain but delays that of a large β-galactosidase domain as a result of kinetic trapping of its unfolded ensemble. We demonstrate that this trapping occurs through a combination of three distinct mechanisms: a decrease in the rate of structural rearrangements within the nascent chain, an increase in the effective exit tunnel length due to folding outside the cradle, and entanglement of the nascent chain with TF. We present evidence that this TF-induced trapping represents a trade-off between promoting co-translational folding and sterically shielding the nascent chain from aberrant cytosolic interactions that could lead to its aggregation or degradation. PMID:22680285

  14. Analyzing complicated protein folding kinetics rapidly by analytical Laplace inversion using a Tikhonov regularization variant.

    PubMed

    Mulligan, Vikram Khipple; Hadley, Kevin Charles; Chakrabartty, Avijit

    2012-02-01

    Kinetic experiments provide much information about protein folding mechanisms. Time-resolved signals are often best described by expressions with many exponential terms, but this hinders the extraction of rate constants by nonlinear least squares (NLS) fitting. Numerical inverse Laplace transformation, which converts a time-resolved dataset into a spectrum of amplitudes as a function of rate constant, allows easy estimation of the rate constants, amplitudes, and number of processes underlying the data. Here, we present a Tikhonov regularization-based method that converts a dataset into a rate spectrum, subject to regularization constraints, without requiring an iterative search of parameter space. This allows more rapid generation of rate spectra as well as analysis of datasets too noisy to process by existing iterative search algorithms. This method's simplicity also permits highly objective, largely automatic analysis with minimal human guidance. We show that this regularization method reproduces results previously obtained by NLS fitting and that it is effective for analyzing datasets too complex for traditional fitting methods. This method's reliability and speed, as well as its potential for objective, model-free analysis, make it extremely useful as a first step in analysis of complicated noisy datasets and an excellent guide for subsequent NLS analysis.

  15. Kinetic folding design of aptazyme-regulated expression devices as riboswitches for metabolic engineering.

    PubMed

    Sparkman-Yager, David; Correa-Rojas, Rodrigo A; Carothers, James M

    2015-01-01

    Recent developments in the fields of synthetic biology and metabolic engineering have opened the doors for the microbial production of biofuels and other valuable organic compounds. There remain, however, significant metabolic hurdles to the production of these compounds in cost-effective quantities. This is due, in part, to mismatches between the metabolic engineer's desire for high yields and the microbe's desire to survive. Many valuable compounds, or the intermediates necessary for their biosynthesis, prove deleterious at the desired production concentrations. One potential solution to these toxicity-related issues is the implementation of nonnative dynamic genetic control mechanisms that sense excessively high concentrations of metabolic intermediates and respond accordingly to alleviate their impact. One potential class of dynamic regulator is the riboswitch: cis-acting RNA elements that regulate the expression of downstream genes based on the presence of an effector molecule. Here, we present combined methods for constructing aptazyme-regulated expression devices (aREDs) through computational cotranscriptional kinetic folding design and experimental validation. These approaches can be used to engineer aREDs within novel genetic contexts for the predictable, dynamic regulation of gene expression in vivo. PMID:25605393

  16. The equilibrium properties and folding kinetics of an all-atom Go model of the Trp-cage.

    PubMed

    Linhananta, Apichart; Boer, Jesse; MacKay, Ian

    2005-03-15

    The ultrafast-folding 20-residue Trp-cage protein is quickly becoming a new benchmark for molecular dynamics studies. Already several all-atom simulations have probed its equilibrium and kinetic properties. In this work an all-atom Go model is used to accurately represent the side-chain packing and native atomic contacts of the Trp-cage. The model reproduces the hallmark thermodynamics cooperativity of small proteins. Folding simulations observe that in the fast-folding dominant pathway, partial alpha-helical structure forms before hydrophobic core collapse. In the slow-folding secondary pathway, partial core collapse occurs before helical structure. The slow-folding rate of the secondary pathway is attributed to the loss of side-chain rotational freedom, due to the early core collapse, which impedes the helix formation. A major finding is the observation of a low-temperature kinetic intermediate stabilized by a salt bridge between residues Asp-9 and Arg-16. Similar observations [R. Zhou, Proc. Natl. Acad. Sci. U.S.A. 100, 13280 (2003)] were reported in a recent study using an all-atom model of the Trp-cage in explicit water, in which the salt-bridge stabilized intermediate was hypothesized to be the origin of the ultrafast-folding mechanism. A theoretical mutation that eliminates the Asp-9-Arg-16 salt bridge, but leaves the residues intact, is performed. Folding simulations of the mutant Trp-cage observe a two-state free-energy landscape with no kinetic intermediate and a significant decrease in the folding rate, in support of the hypothesis.

  17. The initial value problem in Lagrangian drift kinetic theory

    NASA Astrophysics Data System (ADS)

    Burby, J. W.

    2016-06-01

    > Existing high-order variational drift kinetic theories contain unphysical rapidly varying modes that are not seen at low orders. These unphysical modes, which may be rapidly oscillating, damped or growing, are ushered in by a failure of conventional high-order drift kinetic theory to preserve the structure of its parent model's initial value problem. In short, the (infinite dimensional) system phase space is unphysically enlarged in conventional high-order variational drift kinetic theory. I present an alternative, `renormalized' variational approach to drift kinetic theory that manifestly respects the parent model's initial value problem. The basic philosophy underlying this alternate approach is that high-order drift kinetic theory ought to be derived by truncating the all-orders system phase-space Lagrangian instead of the usual `field particle' Lagrangian. For the sake of clarity, this story is told first through the lens of a finite-dimensional toy model of high-order variational drift kinetics; the analogous full-on drift kinetic story is discussed subsequently. The renormalized drift kinetic system, while variational and just as formally accurate as conventional formulations, does not support the troublesome rapidly varying modes.

  18. Using Kinetic Network Models To Probe Non-Native Salt-Bridge Effects on α-Helix Folding.

    PubMed

    Zhou, Guangfeng; Voelz, Vincent A

    2016-02-11

    Salt-bridge interactions play an important role in stabilizing many protein structures, and have been shown to be designable features for protein design. In this work, we study the effects of non-native salt bridges on the folding of a soluble alanine-based peptide (Fs peptide) using extensive all-atom molecular dynamics simulations performed on the Folding@home distributed computing platform. Using Markov State Models, we show how non-native salt-bridges affect the folding kinetics of Fs peptide by perturbing specific conformational states. Furthermore, we present methods for the automatic detection and analysis of such states. These results provide insight into helix folding mechanisms and useful information to guide simulation-based computational protein design.

  19. Analysis of the pH-dependent thermodynamic stability, local motions, and microsecond folding kinetics of carbonmonoxycytochrome c.

    PubMed

    Kumar, Rajesh

    2016-09-15

    This paper analyzes the effect of pH on thermodynamic stability, low-frequency local motions and microsecond folding kinetics of carbonmonoxycytochrome c (Cyt-CO) all across the alkaline pH-unfolding transition of protein. Thermodynamic analysis of urea-induced unfolding transitions of Cyt-CO measured between pH 6 and pH 11.9 reveals that Cyt-CO is maximally stable at pH∼9.5. Dilution of unfolded Cyt-CO into refolding medium forms a native-like compact state (NCO-state), where Fe(2+)-CO interaction persists. Kinetic and thermodynamic parameters measured for slow thermally-driven CO dissociation (NCO→N+CO) and association (N+CO→NCO) reactions between pH 6.5 and pH 13 reveal that the thermal-motions of M80-containing Ω-loop are decreased in subdenaturing limit of alkaline pH. Laser photolysis of Fe(2+)-CO bond in NCO-state triggers the microsecond folding (NCO→N). The microsecond kinetics measured all across the alkaline pH-unfolding transition of Cyt-CO produce rate rollover in the refolding limb of chevron plot, which suggests a glass transition of NCO en route to N. Between pH 7 and pH 11.9, the natural logarithm of the microsecond folding rate varies by < 1.5 units while the natural logarithm of apparent equilibrium constant varies by 11.8 units. This finding indicates that the pH-dependent ionic-interactions greatly affect the global stability of protein but have very small effect on folding kinetics. PMID:27424489

  20. Thermodynamics and kinetics of protein folding on the ribosome: Alteration in energy landscapes, denatured state, and transition state ensembles

    NASA Astrophysics Data System (ADS)

    O'Brien, Edward; Vendruscolo, Michele; Dobson, Christopher

    2010-03-01

    In vitro experiments examining cotranslational folding utilize ribosome-nascent chain complexes (RNCs) in which the nascent chain is stalled at different points of its biosynthesis on the ribosome. We investigate the thermodynamics, kinetics, and structural properties of RNCs containing five different globular and repeat proteins stalled at ten different nascent chain lengths using coarse grained replica exchange simulations. We find that when the proteins are stalled near the ribosome exit tunnel opening they exhibit altered folding coopserativity, quantified by the van't Hoff enthalpy criterion; a significantly altered denatured state ensemble, in terms of Rg and shape parameters (Rg tensor); and the appearance of partially folded intermediates during cotranslation, evidenced by the appearance of a third basin in the free energy profile. These trends are due in part to excluded volume (crowding) interactions between the ribosome and nascent chain. We perform in silico temperature-jump experiments on the RNCs and examine nascent chain folding kinetics and structural changes in the transition state ensemble at various stall lengths.

  1. Simulating the peptide folding kinetic related spectra based on the Markov State Model.

    PubMed

    Song, Jian; Zhuang, Wei

    2014-01-01

    Optical spectroscopic tools are used to monitor protein folding/unfolding dynamics after a fast triggering such as the laser induced temperature jump. These techniques provide new opportunities for comparison between theory and simulations and atom-level understanding protein folding mechanism. However, the direct comparison still face two main challenges: a gap between folding relevant timescales (microseconds or above) and length of molecular dynamics simulations (typically tens to hundreds of nanoseconds), and difficulty in directly calculating spectroscopic observables from simulation configurations. Markov State Model (MSM) approach is one of the most powerful means which can increase simulations timescale up to microsecond or even millisecond. We address progress on modeling infrared and fluorescence spectroscopic signals of temperature jump induced unfolding dynamics for a few small proteins. The harmoniousness between experiment and theoretical can both improve our understanding of protein folding mechanisms and provide direct validation of those theoretical models. PMID:24446363

  2. Efficient Numerical Reconstruction of Protein Folding Kinetics with Partial Path Sampling and Pathlike Variables

    NASA Astrophysics Data System (ADS)

    Juraszek, J.; Saladino, G.; van Erp, T. S.; Gervasio, F. L.

    2013-03-01

    Numerically predicting rate constants of protein folding and other relevant biological events is still a significant challenge. We show that the combination of partial path transition interface sampling with the optimal interfaces and free-energy profiles provided by path collective variables makes the rate calculation for practical biological applications feasible and efficient. This methodology can reproduce the experimental rate constant of Trp-cage miniprotein folding with the same level of accuracy as transition path sampling at a fraction of the cost.

  3. Fold prediction problem: the application of new physical and physicochemical-based features.

    PubMed

    Dehzangi, Abdollah; Phon-Amnuaisuk, Somnuk

    2011-02-01

    One of the most important goals in bioinformatics is the ability to predict tertiary structure of a protein from its amino acid sequence. In this paper, new feature groups based on the physical and physicochemical properties of amino acids (size of the amino acids' side chains, predicted secondary structure based on normalized frequency of β-Strands, Turns, and Reverse Turns) are proposed to tackle this task. The proposed features are extracted using a modified feature extraction method adapted from Dubchak et al. To study the effectiveness of the proposed features and the modified feature extraction method, AdaBoost.M1, Multi Layer Perceptron (MLP), and Support Vector Machine (SVM) that have been commonly and successfully applied to the protein folding problem are employed. Our experimental results show that the new feature groups altogether with the modified feature extraction method are capable of enhancing the protein fold prediction accuracy better than the previous works found in the literature.

  4. A probabilistic approach to the effect of water hydrogen bonds on the kinetics of protein folding and protein denaturation.

    PubMed

    Djikaev, Y S; Ruckenstein, E

    2010-02-26

    Previously, we presented a review of our kinetic models for the nucleation mechanism of protein folding and for the protein thermal denaturation in a barrierless way. A protein was treated as a random heteropolymer consisting of hydrophobic, hydrophilic, and neutral beads. As a crucial idea of the model, an overall potential around the cluster of native residues wherein a residue performs a chaotic motion was considered as the combination of the average dihedral, effective pairwise, and confining potentials. The overall potential as a function of the distance from the cluster has a double well shape. This allowed one to develop kinetic models for the nucleation mechanism of protein folding (NMPF) and barrierless protein denaturation (BPD) by using the mean first passage time analysis. In the original models, however, hydrogen bonding effects were taken into account only indirectly which affected the accuracy of the models because hydrogen bonding does play a crucial role in the folding, stability, and denaturation of proteins. To improve the NMPF and BPD models and explicitly take into account the hydrogen bonding "water-water" and "water-protein residue", we have developed a probabilistic hydrogen bond (PHB) model for the effect of hydrogen bond networks of water molecules around two solute particles (immersed in water) on their interaction, and have then combined the PHB model with the NMPF and BPD models. In this paper, that can be regarded as sequel of our previous review, we analyze the modified NMPF and BPD models that explicitly take into account the effect of water-water hydrogen bonding on these processes. As expected, the application of the modified models to the folding/unfolding of two model proteins (one short, consisting of 124 residues and the other large, consisting of 2500 residues) demonstrate that the hydrogen bond networks play a very important role in the protein folding/unfolding phenomena.

  5. Development of a Fast Microfluidic Mixer for Studies of Protein Folding KineticsFinal Report Cover Page

    SciTech Connect

    Bakajin, O

    2005-02-10

    We designed and fabricated mixing devices that will help us elucidate the mechanisms of protein folding through measurements of folding reaction rates. These devices can be used in studying of other biological systems and are compatible with various spectroscopic observation methods. The project involved development of fabrication processes and setup of a laboratory for assembly and characterization of microfluidic devices, as well as measurements of protein folding kinetics. We produced three variants of the mixer: (1) The ultra fast mixer for Foerster Resonance Energy Transfer measurements (described by Anal. Chem. Article UCRL-JRNL-206676) and MicroTAS Conference Proceedings article (UCRL-JC-153057 ) included in the report; (2) The ultra fast mixer for UV measurements (described by the poster presented at MicroTAS conference (UCRL-POST-207476) included in the report); and (3) The mixer for single molecule measurements (described by the Science article UCRL-JC-153057) included in the report. In these mixers, the channels are narrow, ranging from a few to hundreds of {micro}m, so that the flow is laminar and all of the mixing is achieved through diffusion. Our goal is to develop robust microfluidic mixer with at least 100 times lower consumption rate, shorter dead time and time resolution than commercially available mixers that would be compatible with most commonly used spectroscopic methods. We are also developing mixers that can be used in combination with single molecule spectroscopy. The mixers are used to study kinetics of fast protein folding reactions using bulk fluorescence and single molecule fluorescence resonance energy transfer techniques. Capabilities for microfluidic have been developed at BSNL that will be useful for studies of interactions of DNA with proteins and other projects such as the single molecule detector for detection of low concentration of toxins.

  6. RNA under tension: Folding Landscapes, Kinetic partitioning Mechanism, and Molecular Tensegrity.

    PubMed

    Lin, Jong-Chin; Hyeon, Changbong; Thirumalai, D

    2012-11-19

    Non-coding RNA sequences play a great role in controlling a number of cellular functions, thus raising the need to understand their complex conformational dynamics in quantitative detail. In this perspective, we first show that single molecule pulling when combined with with theory and simulations can be used to quantitatively explore the folding landscape of nucleic acid hairpins, and riboswitches with tertiary interactions. Applications to riboswitches, which are non-coding RNA elements that control gene expression by undergoing dynamical conformational changes in response to binding of metabolites, lead to an organization principle that assembly of RNA is determined by the stability of isolated helices. We also point out the limitations of single molecule pulling experiments, with molecular extension as the only accessible parameter, in extracting key parameters of the folding landscapes of RNA molecules. PMID:23336034

  7. Hg(II) binding to a weakly associated coiled coil nucleates an encoded metalloprotein fold: a kinetic analysis.

    PubMed

    Farrer, Brian T; Pecoraro, Vincent L

    2003-04-01

    A detailed kinetic analysis of metal encapsulation by a de novo-designed protein is described. The kinetic mechanism of Hg(II) encapsulation in the three-stranded coiled coil formed by the peptide CH(3)CO-G LKALEEK CKALEEK LKALEEK G-NH(2) (Baby L9C) is derived by global analysis. The mechanism involves rapid initial collapse of two peptides by Hg(II) forming Hg(Baby L9C(-H))(2) with a linear thiolato Hg(II) bound to the cysteine sulfur atoms. Here, Baby L9C(-H) denotes Baby L9C with the cysteine thiol deprotonated. Addition of the third peptide, forming the three-stranded coiled coil, is the rate-determining step and results in an intermediate state involving two separate species. One of the species, termed the properly folded intermediate, undergoes rapid deprotonation of the third cysteine thiol, yielding the desired three-stranded coiled coil with an encapsulated trigonal thiolato Hg(II). The other species, termed the misfolded intermediate, rearranges in an experimentally distinguishable step to the properly folded intermediate. The order of the reaction involving the addition of the third peptide with respect to the concentration of Baby L9C indicates that addition of the third helix only occurs through reaction of Hg(Baby L9C(-H))(2) and Baby L9C that is unassociated with a coiled coil. Temperature dependence of the reaction afforded activation parameters for both the addition of the third helix (deltaH = 20(2) kcalmol; deltaS= 40(5) calmol K) and the rearrangement of the misfolded intermediate steps (deltaH = 23(2) kcalmol; deltaS= 27(5) calmol K). The mechanism is discussed with regard to metalloprotein folding and metalloprotein design.

  8. Diffusion-collision of foldons elucidates the kinetic effects of point mutations and suggests control strategies of the folding process of helical proteins.

    PubMed

    Capriotti, Emidio; Compiani, Mario

    2006-07-01

    In this article we use mutation studies as a benchmark for a minimal model of the folding process of helical proteins. The model ascribes a pivotal role to the collisional dynamics of a few crucial residues (foldons) and predicts the folding rates by exploiting information drawn from the protein sequence. We show that our model rationalizes the effects of point mutations on the kinetics of folding. The folding times of two proteins and their mutants are predicted. Stability and location of foldons have a critical role as the determinants of protein folding. This allows us to elucidate two main mechanisms for the kinetic effects of mutations. First, it turns out that the mutations eliciting the most notable effects alter protein stability through stabilization or destabilization of the foldons. Secondly, the folding rate is affected via a modification of the foldon topology by those mutations that lead to the birth or death of foldons. The few mispredicted folding rates of some mutants hint at the limits of the current version of the folding model proposed in the present article. The performance of our folding model declines in case the mutated residues are subject to strong long-range forces. That foldons are the critical targets of mutation studies has notable implications for design strategies and is of particular interest to address the issue of the kinetic regulation of single proteins in the general context of the overall dynamics of the interactome.

  9. Temperature dependence of the folding and unfolding kinetics of the GCN4 leucine zipper via 13C(alpha)-NMR.

    PubMed Central

    Holtzer, M E; Bretthorst, G L; d'Avignon, D A; Angeletti, R H; Mints, L; Holtzer, A

    2001-01-01

    Studies by one-dimensional NMR are reported on the interconversion of folded and unfolded forms of the GCN4 leucine zipper in neutral saline buffer. The peptide bears 99% 13C(alpha) labels at three sites: V9, L12, and G31. Time-domain 13C(alpha)-NMR spectra are interpreted by global Bayesian lineshape analysis to extract the rate constants for both unfolding and folding as functions of temperature in the range 47-71 degrees C. The data are well fit by the assumption that the same rate constants apply at each labeled site, confirming that only two conformational states need be considered. Results show that 1) both processes require a free energy of activation; 2) unfolding is kinetically enthalpy-opposed and entropy-driven, while folding is the opposite; and 3) the transition state dimer ensemble averages approximately 40% helical. The activation parameters for unfolding, derived from NMR data at the elevated temperatures where both conformations are populated, lead to estimates of the rate constant at low temperatures (5-15 degrees C) that agree with extant values determined by stopped-flow CD via dilution from denaturing media. However, the corresponding estimated values for the folding rate constant are larger by two to three orders of magnitude than those obtained by stopped flow. We propose that this apparent disagreement is caused by the necessity, in the stopped-flow experiment, for initiation of new helices as the highly denaturant-unfolded molecule adjusts to the newly created benign solvent conditions. This must reduce the success rate of collisions in producing the folded molecule. In the NMR determinations, however, the unfolded chains always have a small, but essential, helix content that makes such initiation unnecessary. Support for this hypothesis is adduced from recent extant experiments on the helix-coil transition in single-chain helical peptides and from demonstration that the folding rate constants for coiled coils, as obtained by stopped flow

  10. Kinetic partitioning mechanism governs the folding of the third FnIII domain of tenascin-C: evidence at the single-molecule level.

    PubMed

    Peng, Qing; Fang, Jie; Wang, Meijia; Li, Hongbin

    2011-09-30

    Statistical mechanics and molecular dynamics simulations proposed that the folding of proteins can follow multiple parallel pathways on a rugged energy landscape from unfolded state en route to their folded native states. Kinetic partitioning mechanism is one of the possible mechanisms underlying such complex folding dynamics. Here, we use single-molecule atomic force microscopy technique to directly probe the multiplicity of the folding pathways of the third fibronectin type III domain from the extracellular matrix protein tenascin-C (TNfn3). By stretching individual (TNfn3)(8) molecules, we forced TNfn3 domains to undergo mechanical unfolding and refolding cycles, allowing us to directly observe the folding pathways of TNfn3. We found that, after being mechanically unraveled and then relaxed to zero force, TNfn3 follows multiple parallel pathways to fold into their native states. The majority of TNfn3 fold into the native state in a simple two-state fashion, while a small percentage of TNfn3 were found to be trapped into kinetically stable folding intermediate states with well-defined three-dimensional structures. Furthermore, the folding of TNfn3 was also influenced by its neighboring TNfn3 domains. Complex misfolded states of TNfn3 were observed, possibly due to the formation of domain-swapped dimeric structures. Our studies revealed the ruggedness of the folding energy landscape of TNfn3 and provided direct experimental evidence that the folding dynamics of TNfn3 are governed by the kinetic partitioning mechanism. Our results demonstrated the unique capability of single-molecule AFM to probe the folding dynamics of proteins at the single-molecule level.

  11. Protein folding, protein structure and the origin of life: Theoretical methods and solutions of dynamical problems

    NASA Technical Reports Server (NTRS)

    Weaver, D. L.

    1982-01-01

    Theoretical methods and solutions of the dynamics of protein folding, protein aggregation, protein structure, and the origin of life are discussed. The elements of a dynamic model representing the initial stages of protein folding are presented. The calculation and experimental determination of the model parameters are discussed. The use of computer simulation for modeling protein folding is considered.

  12. Multiphase Simulated Annealing Based on Boltzmann and Bose-Einstein Distribution Applied to Protein Folding Problem.

    PubMed

    Frausto-Solis, Juan; Liñán-García, Ernesto; Sánchez-Hernández, Juan Paulo; González-Barbosa, J Javier; González-Flores, Carlos; Castilla-Valdez, Guadalupe

    2016-01-01

    A new hybrid Multiphase Simulated Annealing Algorithm using Boltzmann and Bose-Einstein distributions (MPSABBE) is proposed. MPSABBE was designed for solving the Protein Folding Problem (PFP) instances. This new approach has four phases: (i) Multiquenching Phase (MQP), (ii) Boltzmann Annealing Phase (BAP), (iii) Bose-Einstein Annealing Phase (BEAP), and (iv) Dynamical Equilibrium Phase (DEP). BAP and BEAP are simulated annealing searching procedures based on Boltzmann and Bose-Einstein distributions, respectively. DEP is also a simulated annealing search procedure, which is applied at the final temperature of the fourth phase, which can be seen as a second Bose-Einstein phase. MQP is a search process that ranges from extremely high to high temperatures, applying a very fast cooling process, and is not very restrictive to accept new solutions. However, BAP and BEAP range from high to low and from low to very low temperatures, respectively. They are more restrictive for accepting new solutions. DEP uses a particular heuristic to detect the stochastic equilibrium by applying a least squares method during its execution. MPSABBE parameters are tuned with an analytical method, which considers the maximal and minimal deterioration of problem instances. MPSABBE was tested with several instances of PFP, showing that the use of both distributions is better than using only the Boltzmann distribution on the classical SA.

  13. Multiphase Simulated Annealing Based on Boltzmann and Bose-Einstein Distribution Applied to Protein Folding Problem

    PubMed Central

    Liñán-García, Ernesto; Sánchez-Hernández, Juan Paulo; González-Barbosa, J. Javier; González-Flores, Carlos

    2016-01-01

    A new hybrid Multiphase Simulated Annealing Algorithm using Boltzmann and Bose-Einstein distributions (MPSABBE) is proposed. MPSABBE was designed for solving the Protein Folding Problem (PFP) instances. This new approach has four phases: (i) Multiquenching Phase (MQP), (ii) Boltzmann Annealing Phase (BAP), (iii) Bose-Einstein Annealing Phase (BEAP), and (iv) Dynamical Equilibrium Phase (DEP). BAP and BEAP are simulated annealing searching procedures based on Boltzmann and Bose-Einstein distributions, respectively. DEP is also a simulated annealing search procedure, which is applied at the final temperature of the fourth phase, which can be seen as a second Bose-Einstein phase. MQP is a search process that ranges from extremely high to high temperatures, applying a very fast cooling process, and is not very restrictive to accept new solutions. However, BAP and BEAP range from high to low and from low to very low temperatures, respectively. They are more restrictive for accepting new solutions. DEP uses a particular heuristic to detect the stochastic equilibrium by applying a least squares method during its execution. MPSABBE parameters are tuned with an analytical method, which considers the maximal and minimal deterioration of problem instances. MPSABBE was tested with several instances of PFP, showing that the use of both distributions is better than using only the Boltzmann distribution on the classical SA. PMID:27413369

  14. Multiphase Simulated Annealing Based on Boltzmann and Bose-Einstein Distribution Applied to Protein Folding Problem.

    PubMed

    Frausto-Solis, Juan; Liñán-García, Ernesto; Sánchez-Hernández, Juan Paulo; González-Barbosa, J Javier; González-Flores, Carlos; Castilla-Valdez, Guadalupe

    2016-01-01

    A new hybrid Multiphase Simulated Annealing Algorithm using Boltzmann and Bose-Einstein distributions (MPSABBE) is proposed. MPSABBE was designed for solving the Protein Folding Problem (PFP) instances. This new approach has four phases: (i) Multiquenching Phase (MQP), (ii) Boltzmann Annealing Phase (BAP), (iii) Bose-Einstein Annealing Phase (BEAP), and (iv) Dynamical Equilibrium Phase (DEP). BAP and BEAP are simulated annealing searching procedures based on Boltzmann and Bose-Einstein distributions, respectively. DEP is also a simulated annealing search procedure, which is applied at the final temperature of the fourth phase, which can be seen as a second Bose-Einstein phase. MQP is a search process that ranges from extremely high to high temperatures, applying a very fast cooling process, and is not very restrictive to accept new solutions. However, BAP and BEAP range from high to low and from low to very low temperatures, respectively. They are more restrictive for accepting new solutions. DEP uses a particular heuristic to detect the stochastic equilibrium by applying a least squares method during its execution. MPSABBE parameters are tuned with an analytical method, which considers the maximal and minimal deterioration of problem instances. MPSABBE was tested with several instances of PFP, showing that the use of both distributions is better than using only the Boltzmann distribution on the classical SA. PMID:27413369

  15. A 3-fold "butterfly valve" in command of the encapsulation's kinetic stability. Molecular baskets at work.

    PubMed

    Wang, Bao-Yu; Bao, Xiaoguang; Yan, Zhiqing; Maslak, Veselin; Hadad, Christopher M; Badjić, Jovica D

    2008-11-12

    Molecular basket 1, composed of a semirigid tris-norbornadiene framework and three revolving pyridine-based gates at the rim, has been built to "dynamically" enclose space and as such regulate molecular encapsulation. The gates were shown to fold via intramolecular hydrogen bonding and thereby form a C3nu symmetrical receptor: the 1H NMR resonance for the amide N-H protons of the pyridine gates appeared downfield (delta= 10.98 ppm), and the N-H vibrational stretch (IR) was observed at 3176 cm(-1). Accordingly, density functional theory (DFT, B3LYP) investigations revealed for the closed conformers of 1 to be energetically the most stable and dominant. The gearing of the pyridine "gates", about their axis, led to the interconversion of two dynamic enantiomers 1A and 1B comprising the clockwise and counterclockwise seam of intramolecular hydrogen bonds. Dynamic 1H NMR spectroscopic measurements and line-shape simulations suggested that the energy barrier of 10.0 kcal/mol (DeltaG++(A/B), 298 K) is required for the 1A/B interconversion, when CCl4 occupies the cavity of 1. Likewise, the activation free energy for CCl4 departing the basket was found to be 13.1 kcal/mol (DeltaG++, 298 K), whereas the thermodynamic stability of 1:CCl4 complex was -2.7 kcal/mol (DeltaGdegrees, 298 K). In view of that, CCl4 (but also (CH3)3CBr) was proposed to escape from, and a molecule of solvent to enter, the basket when the gates rotate about their axis: the exit of CCl4 requires the activation energy of 12.7 kcal/mol (DeltaG++(A/B) + DeltaGdegrees), similar to the experimentally found 13.1 kcal/mol (DeltaG++). PMID:18937455

  16. Constructive methods of invariant manifolds for kinetic problems

    NASA Astrophysics Data System (ADS)

    Gorban, Alexander N.; Karlin, Iliya V.; Zinovyev, Andrei Yu.

    2004-06-01

    The concept of the slow invariant manifold is recognized as the central idea underpinning a transition from micro to macro and model reduction in kinetic theories. We present the Constructive Methods of Invariant Manifolds for model reduction in physical and chemical kinetics, developed during last two decades. The physical problem of reduced description is studied in the most general form as a problem of constructing the slow invariant manifold. The invariance conditions are formulated as the differential equation for a manifold immersed in the phase space ( the invariance equation). The equation of motion for immersed manifolds is obtained ( the film extension of the dynamics). Invariant manifolds are fixed points for this equation, and slow invariant manifolds are Lyapunov stable fixed points, thus slowness is presented as stability. A collection of methods to derive analytically and to compute numerically the slow invariant manifolds is presented. Among them, iteration methods based on incomplete linearization, relaxation method and the method of invariant grids are developed. The systematic use of thermodynamics structures and of the quasi-chemical representation allow to construct approximations which are in concordance with physical restrictions. The following examples of applications are presented: nonperturbative deviation of physically consistent hydrodynamics from the Boltzmann equation and from the reversible dynamics, for Knudsen numbers Kn∼1; construction of the moment equations for nonequilibrium media and their dynamical correction (instead of extension of list of variables) to gain more accuracy in description of highly nonequilibrium flows; determination of molecules dimension (as diameters of equivalent hard spheres) from experimental viscosity data; model reduction in chemical kinetics; derivation and numerical implementation of constitutive equations for polymeric fluids; the limits of macroscopic description for polymer molecules, etc.

  17. Self-catalyzed growth of S layers via an amorphous-to-crystalline transition limited by folding kinetics

    PubMed Central

    Chung, Sungwook; Shin, Seong-Ho; Bertozzi, Carolyn R.; De Yoreo, James J.

    2010-01-01

    The importance of nonclassical, multistage crystallization pathways is increasingly evident from theoretical studies on colloidal systems and experimental investigations of proteins and biomineral phases. Although theoretical predictions suggest that proteins follow these pathways as a result of fluctuations that create unstable dense-liquid states, microscopic studies indicate these states are long-lived. Using in situ atomic force microscopy to follow 2D assembly of S-layer proteins on supported lipid bilayers, we have obtained a molecular-scale picture of multistage protein crystallization that reveals the importance of conformational transformations in directing the pathway of assembly. We find that monomers with an extended conformation first form a mobile adsorbed phase, from which they condense into amorphous clusters. These clusters undergo a phase transition through S-layer folding into crystalline clusters composed of compact tetramers. Growth then proceeds by formation of new tetramers exclusively at cluster edges, implying tetramer formation is autocatalytic. Analysis of the growth kinetics leads to a quantitative model in which tetramer creation is rate limiting. However, the estimated barrier is much smaller than expected for folding of isolated S-layer proteins, suggesting an energetic rationale for this multistage pathway. PMID:20823255

  18. Improved genetic algorithm for the protein folding problem by use of a Cartesian combination operator.

    PubMed Central

    Rabow, A. A.; Scheraga, H. A.

    1996-01-01

    We have devised a Cartesian combination operator and coding scheme for improving the performance of genetic algorithms applied to the protein folding problem. The genetic coding consists of the C alpha Cartesian coordinates of the protein chain. The recombination of the genes of the parents is accomplished by: (1) a rigid superposition of one parent chain on the other, to make the relation of Cartesian coordinates meaningful, then, (2) the chains of the children are formed through a linear combination of the coordinates of their parents. The children produced with this Cartesian combination operator scheme have similar topology and retain the long-range contacts of their parents. The new scheme is significantly more efficient than the standard genetic algorithm methods for locating low-energy conformations of proteins. The considerable superiority of genetic algorithms over Monte Carlo optimization methods is also demonstrated. We have also devised a new dynamic programming lattice fitting procedure for use with the Cartesian combination operator method. The procedure finds excellent fits of real-space chains to the lattice while satisfying bond-length, bond-angle, and overlap constraints. PMID:8880904

  19. Extreme Folding

    NASA Astrophysics Data System (ADS)

    Demaine, Erik

    2012-02-01

    Our understanding of the mathematics and algorithms behind paper folding, and geometric folding in general, has increased dramatically over the past several years. These developments have found a surprisingly broad range of applications. In the art of origami, it has helped spur the technical origami revolution. In engineering and science, it has helped solve problems in areas such as manufacturing, robotics, graphics, and protein folding. On the recreational side, it has led to new kinds of folding puzzles and magic. I will give an overview of the mathematics and algorithms of folding, with a focus on new mathematics and sculpture.

  20. Kinetics of α-Globin Binding to α-Hemoglobin Stabilizing Protein (AHSP) Indicate Preferential Stabilization of Hemichrome Folding Intermediate*

    PubMed Central

    Mollan, Todd L.; Khandros, Eugene; Weiss, Mitchell J.; Olson, John S.

    2012-01-01

    Human α-hemoglobin stabilizing protein (AHSP) is a conserved mammalian erythroid protein that facilitates the production of Hemoglobin A by stabilizing free α-globin. AHSP rapidly binds to ferrous α with association (k′AHSP) and dissociation (kAHSP) rate constants of ≈10 μm−1 s−1 and 0.2 s−1, respectively, at pH 7.4 at 22 °C. A small slow phase was observed when AHSP binds to excess ferrous αCO. This slow phase appears to be due to cis to trans prolyl isomerization of the Asp29-Pro30 peptide bond in wild-type AHSP because it was absent when αCO was mixed with P30A and P30W AHSP, which are fixed in the trans conformation. This slow phase was also absent when met(Fe3+)-α reacted with wild-type AHSP, suggesting that met-α is capable of rapidly binding to either Pro30 conformer. Both wild-type and Pro30-substituted AHSPs drive the formation of a met-α hemichrome conformation following binding to either met- or oxy(Fe2+)-α. The dissociation rate of the met-α·AHSP complex (kAHSP ≈ 0.002 s−1) is ∼100-fold slower than that for ferrous α·AHSP complexes, resulting in a much higher affinity of AHSP for met-α. Thus, in vivo, AHSP acts as a molecular chaperone by rapidly binding and stabilizing met-α hemichrome folding intermediates. The low rate of met-α dissociation also allows AHSP to have a quality control function by kinetically trapping ferric α and preventing its incorporation into less stable mixed valence Hemoglobin A tetramers. Reduction of AHSP-bound met-α allows more rapid release to β subunits to form stable fully, reduced hemoglobin dimers and tetramers. PMID:22298770

  1. Kinetic and thermodynamic analysis of the conformational folding process of SS-reduced bovine pancreatic ribonuclease A using a selenoxide reagent with high oxidizing ability.

    PubMed

    Arai, Kenta; Kumakura, Fumio; Iwaoka, Michio

    2012-01-01

    Redox-coupled folding pathways of bovine pancreatic ribonuclease A (RNase A) with four intramolecular disulfide (SS) bonds comprise three phases: (I) SS formation to generate partially oxidized intermediate ensembles with no rigid folded structure; (II) SS rearrangement from the three SS intermediate ensemble (3S) to the des intermediates having three native SS linkages; (III) final oxidation of the last native SS linkage to generate native RNase A. We previously demonstrated that DHS(ox), a water-soluble selenoxide reagent for rapid and quantitative SS formation, allows clear separation of the three folding phases. In this study, the main conformational folding phase (phase II) has been extensively analyzed at pH 8.0 under a wide range of temperatures (5-45 °C), and thermodynamic and kinetic parameters for the four des intermediates were determined. The free-energy differences (ΔG) as a function of temperature suggested that the each SS linkage has different thermodynamic and kinetic roles in stability of the native structure. On the other hand, comparison of the rate constants and the activation energies for 3S → des with those reported for the conformational folding of SS-intact RNase A suggested that unfolded des species (desU) having three native SS linkages but not yet being folded are involved in very small amounts (<1%) in the 3S intermediate ensemble and the desU species would gain the native-like structures via X-Pro isomerization like SS-intact RNase A. It was revealed that DHS(ox) is useful for kinetic and thermodynamic analysis of the conformational folding process on the oxidative folding pathways of SS-reduced proteins.

  2. The equilibrium properties and folding kinetics of an all-atom Go xAF model of the Trp-cage

    NASA Astrophysics Data System (ADS)

    Linhananta, Apichart; Boer, Jesse; MacKay, Ian

    2005-03-01

    The ultrafast-folding 20-residue Trp-cage protein is quickly becoming a new benchmark for molecular dynamics studies. Already several all-atom simulations have probed its equilibrium and kinetic properties. In this work an all-atom Go ¯ model is used to accurately represent the side-chain packing and native atomic contacts of the Trp-cage. The model reproduces the hallmark thermodynamics cooperativity of small proteins. Folding simulations observe that in the fast-folding dominant pathway, partial α-helical structure forms before hydrophobic core collapse. In the slow-folding secondary pathway, partial core collapse occurs before helical structure. The slow-folding rate of the secondary pathway is attributed to the loss of side-chain rotational freedom, due to the early core collapse, which impedes the helix formation. A major finding is the observation of a low-temperature kinetic intermediate stabilized by a salt bridge between residues Asp-9 and Arg-16. Similar observations [R. Zhou, Proc. Natl. Acad. Sci. U.S.A. 100, 13280 (2003)] were reported in a recent study using an all-atom model of the Trp-cage in explicit water, in which the salt-bridge stabilized intermediate was hypothesized to be the origin of the ultrafast-folding mechanism. A theoretical mutation that eliminates the Asp-9-Arg-16 salt bridge, but leaves the residues intact, is performed. Folding simulations of the mutant Trp-cage observe a two-state free-energy landscape with no kinetic intermediate and a significant decrease in the folding rate, in support of the hypothesis.

  3. Comparative analysis of the folding dynamics and kinetics of an engineered knotted protein and its variants derived from HP0242 of Helicobacter pylori

    NASA Astrophysics Data System (ADS)

    Wang, Liang-Wei; Liu, Yu-Nan; Lyu, Ping-Chiang; Jackson, Sophie E.; Hsu, Shang-Te Danny

    2015-09-01

    Understanding the mechanism by which a polypeptide chain thread itself spontaneously to attain a knotted conformation has been a major challenge in the field of protein folding. HP0242 is a homodimeric protein from Helicobacter pylori with intertwined helices to form a unique pseudo-knotted folding topology. A tandem HP0242 repeat has been constructed to become the first engineered trefoil-knotted protein. Its small size renders it a model system for computational analyses to examine its folding and knotting pathways. Here we report a multi-parametric study on the folding stability and kinetics of a library of HP0242 variants, including the trefoil-knotted tandem HP0242 repeat, using far-UV circular dichroism and fluorescence spectroscopy. Equilibrium chemical denaturation of HP0242 variants shows the presence of highly populated dimeric and structurally heterogeneous folding intermediates. Such equilibrium folding intermediates retain significant amount of helical structures except those at the N- and C-terminal regions in the native structure. Stopped-flow fluorescence measurements of HP0242 variants show that spontaneous refolding into knotted structures can be achieved within seconds, which is several orders of magnitude faster than previously observed for other knotted proteins. Nevertheless, the complex chevron plots indicate that HP0242 variants are prone to misfold into kinetic traps, leading to severely rolled-over refolding arms. The experimental observations are in general agreement with the previously reported molecular dynamics simulations. Based on our results, kinetic folding pathways are proposed to qualitatively describe the complex folding processes of HP0242 variants.

  4. Kinetic Transition Networks for the Thomson Problem and Smale's Seventh Problem

    NASA Astrophysics Data System (ADS)

    Mehta, Dhagash; Chen, Jianxu; Chen, Danny Z.; Kusumaatmaja, Halim; Wales, David J.

    2016-07-01

    The Thomson problem, arrangement of identical charges on the surface of a sphere, has found many applications in physics, chemistry and biology. Here, we show that the energy landscape of the Thomson problem for N particles with N =132 , 135, 138, 141, 144, 147, and 150 is single funneled, characteristic of a structure-seeking organization where the global minimum is easily accessible. Algorithmically, constructing starting points close to the global minimum of such a potential with spherical constraints is one of Smale's 18 unsolved problems in mathematics for the 21st century because it is important in the solution of univariate and bivariate random polynomial equations. By analyzing the kinetic transition networks, we show that a randomly chosen minimum is, in fact, always "close" to the global minimum in terms of the number of transition states that separate them, a characteristic of small world networks.

  5. Kinetic Transition Networks for the Thomson Problem and Smale's Seventh Problem.

    PubMed

    Mehta, Dhagash; Chen, Jianxu; Chen, Danny Z; Kusumaatmaja, Halim; Wales, David J

    2016-07-01

    The Thomson problem, arrangement of identical charges on the surface of a sphere, has found many applications in physics, chemistry and biology. Here, we show that the energy landscape of the Thomson problem for N particles with N=132, 135, 138, 141, 144, 147, and 150 is single funneled, characteristic of a structure-seeking organization where the global minimum is easily accessible. Algorithmically, constructing starting points close to the global minimum of such a potential with spherical constraints is one of Smale's 18 unsolved problems in mathematics for the 21st century because it is important in the solution of univariate and bivariate random polynomial equations. By analyzing the kinetic transition networks, we show that a randomly chosen minimum is, in fact, always "close" to the global minimum in terms of the number of transition states that separate them, a characteristic of small world networks. PMID:27447530

  6. An ant colony optimisation algorithm for the 2D and 3D hydrophobic polar protein folding problem

    PubMed Central

    Shmygelska, Alena; Hoos, Holger H

    2005-01-01

    Background The protein folding problem is a fundamental problems in computational molecular biology and biochemical physics. Various optimisation methods have been applied to formulations of the ab-initio folding problem that are based on reduced models of protein structure, including Monte Carlo methods, Evolutionary Algorithms, Tabu Search and hybrid approaches. In our work, we have introduced an ant colony optimisation (ACO) algorithm to address the non-deterministic polynomial-time hard (NP-hard) combinatorial problem of predicting a protein's conformation from its amino acid sequence under a widely studied, conceptually simple model – the 2-dimensional (2D) and 3-dimensional (3D) hydrophobic-polar (HP) model. Results We present an improvement of our previous ACO algorithm for the 2D HP model and its extension to the 3D HP model. We show that this new algorithm, dubbed ACO-HPPFP-3, performs better than previous state-of-the-art algorithms on sequences whose native conformations do not contain structural nuclei (parts of the native fold that predominantly consist of local interactions) at the ends, but rather in the middle of the sequence, and that it generally finds a more diverse set of native conformations. Conclusions The application of ACO to this bioinformatics problem compares favourably with specialised, state-of-the-art methods for the 2D and 3D HP protein folding problem; our empirical results indicate that our rather simple ACO algorithm scales worse with sequence length but usually finds a more diverse ensemble of native states. Therefore the development of ACO algorithms for more complex and realistic models of protein structure holds significant promise. PMID:15710037

  7. Structural analysis of kinetic folding intermediates for a TIM barrel protein, indole-3-glycerol phosphate synthase, by hydrogen exchange mass spectrometry and Gō model simulation.

    PubMed

    Gu, Zhenyu; Rao, Maithreyi K; Forsyth, William R; Finke, John M; Matthews, C Robert

    2007-11-23

    The structures of partially folded states appearing during the folding of a (betaalpha)(8) TIM barrel protein, the indole-3-glycerol phosphate synthase from Sulfolobus solfataricus (sIGPS), was assessed by hydrogen exchange mass spectrometry (HX-MS) and Gō model simulations. HX-MS analysis of the peptic peptides derived from the pulse-labeled product of the sub-millisecond folding reaction from the urea-denatured state revealed strong protection in the (betaalpha)(4) region, modest protection in the neighboring (betaalpha)(1-3) and (betaalpha)(5)beta(6) segments and no significant protection in the remaining N and C-terminal segments. These results demonstrate that this species is not a collapsed form of the unfolded state under native-favoring conditions nor is it the native state formed via fast-track folding. However, the striking contrast of these results with the strong protection observed in the (betaalpha)(2-5)beta(6) region after 5 s of folding demonstrates that these species represent kinetically distinct folding intermediates that are not identical as previously thought. A re-examination of the kinetic folding mechanism by chevron analysis of fluorescence data confirmed distinct roles for these two species: the burst-phase intermediate is predicted to be a misfolded, off-pathway intermediate, while the subsequent 5 s intermediate corresponds to an on-pathway equilibrium intermediate. Comparison with the predictions using a C(alpha) Gō model simulation of the kinetic folding reaction for sIGPS shows good agreement with the core of the structure offering protection against exchange in the on-pathway intermediate(s). Because the native-centric Gō model simulations do not explicitly include sequence-specific information, the simulation results support the hypothesis that the topology of TIM barrel proteins is a primary determinant of the folding free energy surface for the productive folding reaction. The early misfolding reaction must involve aspects of

  8. Kinetic and thermodynamic framework for P4-P6 RNA reveals tertiary motif modularity and modulation of the folding preferred pathway.

    PubMed

    Bisaria, Namita; Greenfeld, Max; Limouse, Charles; Pavlichin, Dmitri S; Mabuchi, Hideo; Herschlag, Daniel

    2016-08-23

    The past decade has seen a wealth of 3D structural information about complex structured RNAs and identification of functional intermediates. Nevertheless, developing a complete and predictive understanding of the folding and function of these RNAs in biology will require connection of individual rate and equilibrium constants to structural changes that occur in individual folding steps and further relating these steps to the properties and behavior of isolated, simplified systems. To accomplish these goals we used the considerable structural knowledge of the folded, unfolded, and intermediate states of P4-P6 RNA. We enumerated structural states and possible folding transitions and determined rate and equilibrium constants for the transitions between these states using single-molecule FRET with a series of mutant P4-P6 variants. Comparisons with simplified constructs containing an isolated tertiary contact suggest that a given tertiary interaction has a stereotyped rate for breaking that may help identify structural transitions within complex RNAs and simplify the prediction of folding kinetics and thermodynamics for structured RNAs from their parts. The preferred folding pathway involves initial formation of the proximal tertiary contact. However, this preference was only ∼10 fold and could be reversed by a single point mutation, indicating that a model akin to a protein-folding contact order model will not suffice to describe RNA folding. Instead, our results suggest a strong analogy with a modified RNA diffusion-collision model in which tertiary elements within preformed secondary structures collide, with the success of these collisions dependent on whether the tertiary elements are in their rare binding-competent conformations. PMID:27493222

  9. Single-Molecule Fluorescence Resonance Energy Transfer Studies of the Human Telomerase RNA Pseudoknot: Temperature-/Urea-Dependent Folding Kinetics and Thermodynamics

    PubMed Central

    2015-01-01

    The ribonucleoprotein telomerase is an RNA-dependent DNA polymerase that catalyzes the repetitive addition of a short, species-specific, DNA sequence to the ends of linear eukaryotic chromosomes. The single RNA component of telomerase contains both the template sequence for DNA synthesis and a functionally critical pseudoknot motif, which can also exist as a less stable hairpin. Here we use a minimal version of the human telomerase RNA pseudoknot to study this hairpin–pseudoknot structural equilibrium using temperature-controlled single-molecule fluorescence resonance energy transfer (smFRET) experiments. The urea dependence of these experiments aids in determination of the folding kinetics and thermodynamics. The wild-type pseudoknot behavior is compared and contrasted to a mutant pseudoknot sequence implicated in a genetic disorder–dyskeratosis congenita. These findings clearly identify that this 2nt noncomplementary mutation destabilizes the folding of the wild-type pseudoknot by substantially reducing the folding rate constant (≈ 400-fold) while only nominally increasing the unfolding rate constant (≈ 5-fold). Furthermore, the urea dependence of the equilibrium and rate constants is used to develop a free energy landscape for this unimolecular equilibrium and propose details about the structure of the transition state. Finally, the urea-dependent folding experiments provide valuable physical insights into the mechanism for destabilization of RNA pseudoknots by such chemical denaturants. PMID:24617561

  10. Rapid three-dimensional microfluidic mixer for high viscosity solutions to unravel earlier folding kinetics of G-quadruplex under molecular crowding conditions.

    PubMed

    Liu, Chao; Li, Ying; Li, Yiwei; Chen, Peng; Feng, Xiaojun; Du, Wei; Liu, Bi-Feng

    2016-03-01

    Rapid mixing of highly viscous solutions is a great challenge, which helps to analyze the reaction kinetics in viscous liquid phase, particularly to discover the folding kinetics of macromolecules under molecular crowding conditions mimicking the conditions inside cells. Here, we demonstrated a novel microfluidic mixer based on Dean flows with three-dimensional (3D) microchannel configuration for fast mixing of high-viscosity fluids. The main structure contained three consecutive subunits, each consisting of a "U"-type channel followed by a chamber with different width and height. Thus, the two solutions injected from the two inlets would undergo a mixing in the first "U"-type channel due to the Dean flow effect, and simultaneous vortices expansions in both horizontal and vertical directions in the following chamber. Numerical simulations and experimental characterizations confirmed that the micromixer could achieve a mixing time of 122.4μs for solutions with viscosities about 33.6 times that of pure water. It was the fastest micromixer for high viscosity solutions compared with previous reports. With this highly efficient 3D microfluidic mixer, we further characterized the early folding kinetics of human telomere G-quadruplex under molecular crowding conditions, and unravelled a new folding process within 550μs. PMID:26717836

  11. Rapid three-dimensional microfluidic mixer for high viscosity solutions to unravel earlier folding kinetics of G-quadruplex under molecular crowding conditions.

    PubMed

    Liu, Chao; Li, Ying; Li, Yiwei; Chen, Peng; Feng, Xiaojun; Du, Wei; Liu, Bi-Feng

    2016-01-01

    Rapid mixing of highly viscous solutions is a great challenge, which helps to analyze the reaction kinetics in viscous liquid phase, particularly to discover the folding kinetics of macromolecules under molecular crowding conditions mimicking the conditions inside cells. Here, we demonstrated a novel microfluidic mixer based on Dean flows with three-dimensional (3D) microchannel configuration for fast mixing of high-viscosity fluids. The main structure contained three consecutive subunits, each consisting of a "U"-type channel followed by a chamber with different width and height. Thus, the two solutions injected from the two inlets would undergo a mixing in the first "U"-type channel due to the Dean flow effect, and simultaneous vortices expansions in both horizontal and vertical directions in the following chamber. Numerical simulations and experimental characterizations confirmed that the micromixer could achieve a mixing time of 122.4μs for solutions with viscosities about 33.6 times that of pure water. It was the fastest micromixer for high viscosity solutions compared with previous reports. With this highly efficient 3D microfluidic mixer, we further characterized the early folding kinetics of human telomere G-quadruplex under molecular crowding conditions, and unravelled a new folding process within 550μs.

  12. Network measures for protein folding state discrimination

    PubMed Central

    Menichetti, Giulia; Fariselli, Piero; Remondini, Daniel

    2016-01-01

    Proteins fold using a two-state or multi-state kinetic mechanisms, but up to now there is not a first-principle model to explain this different behavior. We exploit the network properties of protein structures by introducing novel observables to address the problem of classifying the different types of folding kinetics. These observables display a plain physical meaning, in terms of vibrational modes, possible configurations compatible with the native protein structure, and folding cooperativity. The relevance of these observables is supported by a classification performance up to 90%, even with simple classifiers such as discriminant analysis. PMID:27464796

  13. Network measures for protein folding state discrimination.

    PubMed

    Menichetti, Giulia; Fariselli, Piero; Remondini, Daniel

    2016-01-01

    Proteins fold using a two-state or multi-state kinetic mechanisms, but up to now there is not a first-principle model to explain this different behavior. We exploit the network properties of protein structures by introducing novel observables to address the problem of classifying the different types of folding kinetics. These observables display a plain physical meaning, in terms of vibrational modes, possible configurations compatible with the native protein structure, and folding cooperativity. The relevance of these observables is supported by a classification performance up to 90%, even with simple classifiers such as discriminant analysis. PMID:27464796

  14. Influence of Glu/Arg, Asp/Arg, and Glu/Lys Salt Bridges on α-Helical Stability and Folding Kinetics.

    PubMed

    Meuzelaar, Heleen; Vreede, Jocelyne; Woutersen, Sander

    2016-06-01

    Using a combination of ultraviolet circular dichroism, temperature-jump transient-infrared spectroscopy, and molecular dynamics simulations, we investigate the effect of salt bridges between different types of charged amino-acid residue pairs on α-helix folding. We determine the stability and the folding and unfolding rates of 12 alanine-based α-helical peptides, each of which has a nearly identical composition containing three pairs of positively and negatively charged residues (either Glu(-)/Arg(+), Asp(-)/Arg(+), or Glu(-)/Lys(+)). Within each set of peptides, the distance and order of the oppositely charged residues in the peptide sequence differ, such that they have different capabilities of forming salt bridges. Our results indicate that stabilizing salt bridges (in which the interacting residues are spaced and ordered such that they favor helix formation) speed up α-helix formation by up to 50% and slow down the unfolding of the α-helix, whereas salt bridges with an unfavorable geometry have the opposite effect. Comparing the peptides with different types of charge pairs, we observe that salt bridges between side chains of Glu(-) and Arg(+) are most favorable for the speed of folding, probably because of the larger conformational space of the salt-bridging Glu(-)/Arg(+) rotamer pairs compared to Asp(-)/Arg(+) and Glu(-)/Lys(+). We speculate that the observed impact of salt bridges on the folding kinetics might explain why some proteins contain salt bridges that do not stabilize the final, folded conformation. PMID:27276251

  15. Influence of Glu/Arg, Asp/Arg, and Glu/Lys Salt Bridges on α-Helical Stability and Folding Kinetics.

    PubMed

    Meuzelaar, Heleen; Vreede, Jocelyne; Woutersen, Sander

    2016-06-01

    Using a combination of ultraviolet circular dichroism, temperature-jump transient-infrared spectroscopy, and molecular dynamics simulations, we investigate the effect of salt bridges between different types of charged amino-acid residue pairs on α-helix folding. We determine the stability and the folding and unfolding rates of 12 alanine-based α-helical peptides, each of which has a nearly identical composition containing three pairs of positively and negatively charged residues (either Glu(-)/Arg(+), Asp(-)/Arg(+), or Glu(-)/Lys(+)). Within each set of peptides, the distance and order of the oppositely charged residues in the peptide sequence differ, such that they have different capabilities of forming salt bridges. Our results indicate that stabilizing salt bridges (in which the interacting residues are spaced and ordered such that they favor helix formation) speed up α-helix formation by up to 50% and slow down the unfolding of the α-helix, whereas salt bridges with an unfavorable geometry have the opposite effect. Comparing the peptides with different types of charge pairs, we observe that salt bridges between side chains of Glu(-) and Arg(+) are most favorable for the speed of folding, probably because of the larger conformational space of the salt-bridging Glu(-)/Arg(+) rotamer pairs compared to Asp(-)/Arg(+) and Glu(-)/Lys(+). We speculate that the observed impact of salt bridges on the folding kinetics might explain why some proteins contain salt bridges that do not stabilize the final, folded conformation.

  16. A Hooke׳s law-based approach to protein folding rate.

    PubMed

    Ruiz-Blanco, Yasser B; Marrero-Ponce, Yovani; Prieto, Pablo J; Salgado, Jesús; García, Yamila; Sotomayor-Torres, Clivia M

    2015-01-01

    Kinetics is a key aspect of the renowned protein folding problem. Here, we propose a comprehensive approach to folding kinetics where a polypeptide chain is assumed to behave as an elastic material described by the Hooke׳s law. A novel parameter called elastic-folding constant results from our model and is suggested to distinguish between protein with two-state and multi-state folding pathways. A contact-free descriptor, named folding degree, is introduced as a suitable structural feature to study protein-folding kinetics. This approach generalizes the observed correlations between varieties of structural descriptors with the folding rate constant. Additionally several comparisons among structural classes and folding mechanisms were carried out showing the good performance of our model with proteins of different types. The present model constitutes a simple rationale for the structural and energetic factors involved in protein folding kinetics. PMID:25245368

  17. Numerical impact simulation of gradually increased kinetic energy transfer has the potential to break up folded protein structures resulting in cytotoxic brain tissue edema.

    PubMed

    von Holst, Hans; Li, Xiaogai

    2013-07-01

    Although the consequences of traumatic brain injury (TBI) and its treatment have been improved, there is still a substantial lack of understanding the mechanisms. Numerical simulation of the impact can throw further lights on site and mechanism of action. A finite element model of the human head and brain tissue was used to simulate TBI. The consequences of gradually increased kinetic energy transfer was analyzed by evaluating the impact intracranial pressure (ICP), strain level, and their potential influences on binding forces in folded protein structures. The gradually increased kinetic energy was found to have the potential to break apart bonds of Van der Waals in all impacts and hydrogen bonds at simulated impacts from 6 m/s and higher, thereby superseding the energy in folded protein structures. Further, impacts below 6 m/s showed none or very slight increase in impact ICP and strain levels, whereas impacts of 6 m/s or higher showed a gradual increase of the impact ICP and strain levels reaching over 1000 KPa and over 30%, respectively. The present simulation study shows that the free kinetic energy transfer, impact ICP, and strain levels all have the potential to initiate cytotoxic brain tissue edema by unfolding protein structures. The definition of mild, moderate, and severe TBI should thus be looked upon as the same condition and separated only by a gradual severity of impact.

  18. Scanning Single-Molecule Fluorescence Correlation Spectroscopy Enables Kinetics Study of DNA Hairpin Folding with a Time Window from Microseconds to Seconds.

    PubMed

    Bi, Huimin; Yin, Yandong; Pan, Bailong; Li, Geng; Zhao, Xin Sheng

    2016-05-19

    Single-molecule fluorescence measurements have been widely used to explore kinetics and dynamics of biological systems. Among them, single-molecule imaging (SMI) is good at tracking processes slower than tens of milliseconds, whereas fluorescence correlation spectroscopy (FCS) is good at probing processes faster than submilliseconds. However, there is still shortage of simple yet effective single-molecule fluorescence method to cover the time-scale between submilliseconds and tens of milliseconds. To effectively bridge this millisecond gap, we developed a single-molecule fluorescence correlation spectroscopy (smFCS) method that works on surface-immobilized single molecules through surface scanning. We validated it by monitoring the classical DNA hairpin folding process. With a wide time window from microseconds to seconds, the experimental data are well fitted to the two-state folding model. All relevant molecular parameters, including the relative fluorescence brightness, equilibrium constant, and reaction rate constants, were uniquely determined.

  19. A New Folding Kinetic Mechanism for Human Transthyretin and the Influence of the Amyloidogenic V30M Mutation.

    PubMed

    Jesus, Catarina S H; Almeida, Zaida L; Vaz, Daniela C; Faria, Tiago Q; Brito, Rui M M

    2016-08-31

    Protein aggregation into insoluble amyloid fibrils is the hallmark of several neurodegenerative diseases, chief among them Alzheimer's and Parkinson's. Although caused by different proteins, these pathologies share some basic molecular mechanisms with familial amyloidotic polyneuropathy (FAP), a rare hereditary neuropathy caused by amyloid formation and deposition by transthyretin (TTR) in the peripheral and autonomic nervous systems. Among the amyloidogenic TTR mutations known, V30M-TTR is the most common in FAP. TTR amyloidogenesis (ATTR) is triggered by tetramer dissociation, followed by partial unfolding and aggregation of the low conformational stability monomers formed. Thus, tetramer dissociation kinetics, monomer conformational stability and competition between refolding and aggregation pathways do play a critical role in ATTR. Here, we propose a new model to analyze the refolding kinetics of WT-TTR and V30M-TTR, showing that at pH and protein concentrations close to physiological, a two-step mechanism with a unimolecular first step followed by a second-order second step adjusts well to the experimental data. Interestingly, although sharing the same kinetic mechanism, V30M-TTR refolds at a much slower rate than WT-TTR, a feature that may favor the formation of transient species leading to kinetic partition into amyloidogenic pathways and, thus, significantly increasing the probability of amyloid formation in vivo.

  20. A New Folding Kinetic Mechanism for Human Transthyretin and the Influence of the Amyloidogenic V30M Mutation.

    PubMed

    Jesus, Catarina S H; Almeida, Zaida L; Vaz, Daniela C; Faria, Tiago Q; Brito, Rui M M

    2016-01-01

    Protein aggregation into insoluble amyloid fibrils is the hallmark of several neurodegenerative diseases, chief among them Alzheimer's and Parkinson's. Although caused by different proteins, these pathologies share some basic molecular mechanisms with familial amyloidotic polyneuropathy (FAP), a rare hereditary neuropathy caused by amyloid formation and deposition by transthyretin (TTR) in the peripheral and autonomic nervous systems. Among the amyloidogenic TTR mutations known, V30M-TTR is the most common in FAP. TTR amyloidogenesis (ATTR) is triggered by tetramer dissociation, followed by partial unfolding and aggregation of the low conformational stability monomers formed. Thus, tetramer dissociation kinetics, monomer conformational stability and competition between refolding and aggregation pathways do play a critical role in ATTR. Here, we propose a new model to analyze the refolding kinetics of WT-TTR and V30M-TTR, showing that at pH and protein concentrations close to physiological, a two-step mechanism with a unimolecular first step followed by a second-order second step adjusts well to the experimental data. Interestingly, although sharing the same kinetic mechanism, V30M-TTR refolds at a much slower rate than WT-TTR, a feature that may favor the formation of transient species leading to kinetic partition into amyloidogenic pathways and, thus, significantly increasing the probability of amyloid formation in vivo. PMID:27589730

  1. A New Folding Kinetic Mechanism for Human Transthyretin and the Influence of the Amyloidogenic V30M Mutation

    PubMed Central

    Jesus, Catarina S. H.; Almeida, Zaida L.; Vaz, Daniela C.; Faria, Tiago Q.; Brito, Rui M. M.

    2016-01-01

    Protein aggregation into insoluble amyloid fibrils is the hallmark of several neurodegenerative diseases, chief among them Alzheimer’s and Parkinson’s. Although caused by different proteins, these pathologies share some basic molecular mechanisms with familial amyloidotic polyneuropathy (FAP), a rare hereditary neuropathy caused by amyloid formation and deposition by transthyretin (TTR) in the peripheral and autonomic nervous systems. Among the amyloidogenic TTR mutations known, V30M-TTR is the most common in FAP. TTR amyloidogenesis (ATTR) is triggered by tetramer dissociation, followed by partial unfolding and aggregation of the low conformational stability monomers formed. Thus, tetramer dissociation kinetics, monomer conformational stability and competition between refolding and aggregation pathways do play a critical role in ATTR. Here, we propose a new model to analyze the refolding kinetics of WT-TTR and V30M-TTR, showing that at pH and protein concentrations close to physiological, a two-step mechanism with a unimolecular first step followed by a second-order second step adjusts well to the experimental data. Interestingly, although sharing the same kinetic mechanism, V30M-TTR refolds at a much slower rate than WT-TTR, a feature that may favor the formation of transient species leading to kinetic partition into amyloidogenic pathways and, thus, significantly increasing the probability of amyloid formation in vivo. PMID:27589730

  2. Kinetic approach to the evaporation and condensation problem

    NASA Technical Reports Server (NTRS)

    Murakami, M.; Oshima, K.

    1974-01-01

    In the paper, the Boltzmann equation governing the evaporation and condensation phenomena is solved by the Monte Carlo method. Based on the kinetic theory of gas the role of the non-equilibrium Knudsen layer and the growth of the hydrodynamic region outside the layer as time proceeds are simulated. Results show two possible types of transient developments in the vapor phase. The effects of the molecular absorption coefficient of the phase surface are examined. Except in the case of very strong evaporation the kinematic effects of binary collisions among vapor molecules on the mass flux rate are not serious. The limiting case of the quasi-steady evaporation and the maximal value of the evaporation rate are obtained.

  3. The experimental folding landscape of monomeric lactose repressor, a large two-domain protein, involves two kinetic intermediates.

    PubMed

    Wilson, Corey J; Das, Payel; Clementi, Cecilia; Matthews, Kathleen S; Wittung-Stafshede, Pernilla

    2005-10-11

    To probe the experimental folding behavior of a large protein with complex topology, we created a monomeric variant of the lactose repressor protein (MLAc), a well characterized tetrameric protein that regulates transcription of the lac operon. Purified MLAc is folded, fully functional, and binds the inducer isopropyl beta-d-thiogalactoside with the same affinity as wild-type LacI. Equilibrium unfolding of MLAc induced by the chemical denaturant urea is a reversible, apparent two-state process (pH 7.5, 20 degrees C). However, time-resolved experiments demonstrate that unfolding is single-exponential, whereas refolding data indicate two transient intermediates. The data reveal the initial formation of a burst-phase (tau < ms) intermediate that corresponds to approximately 50% of the total secondary-structure content. This step is followed by a rearrangement reaction that is rate-limited by an unfolding process (tau approximately 3 s; pH 7.5, 20 degrees C) and results in a second intermediate. This MLAc intermediate converts to the native structure (tau approximately 30 s; pH 7.5, 20 degrees C). Remarkably, the experimental folding-energy landscape for MLAc is in excellent agreement with theoretical predictions using a simple topology-based C(alpha)-model as presented in a companion article in this issue. PMID:16203983

  4. Multipixel spectral imaging of green fluorescent protein (GFP) in COS-7 cells: folding kinetics and chromophore formation

    NASA Astrophysics Data System (ADS)

    Greenbaum, Lior; Rothmann, Chana; Hanania, Judith; Malik, Zvi

    2000-12-01

    Spectrally resolved imaging of Green fluorescent protein (GFP) expressed in living COS-7 kidney cells distinguished the subcellular localization and demarcated the processes of protein folding and chromophore formation. COS-7 kidney cells were transfected by a plasmid pEGFP-N1 plasmid followed by incubation for 15 hours for gen expression. At different intervals the cells were examined by fluorescence microscopy and analyzed by a spectral imaging system. After 7 hours, GFP was detected in the cytoplasm, concentrated in a localized form. Spectra of the initial GFP evinced several components that belong both tot he typical fluorescent signal as well as to unspecific fingerprints. At 10 and 15 hours, GFP was seen spread in the cytoplasm as well as in the nucleus, and the specific spectra of the GFP were dominant at the later time. The typical GFP spectral fingerprint is the result of protein folding and chromophore formation following internal oxidation reactions. This folding and chromophore formation process, up to final conformation, was detected by spectral imaging as localized in the nucleus rather than in the cytosol. Thus, the method of fluorescence microscopy combined with multiplex spectral imaging demonstrates distinct biochemical pathways leading to GFP conformation.

  5. Distinguishing between sequential and nonsequentially folded proteins: implications for folding and misfolding.

    PubMed Central

    Tsai, C. J.; Maizel, J. V.; Nussinov, R.

    1999-01-01

    We describe here an algorithm for distinguishing sequential from nonsequentially folding proteins. Several experiments have recently suggested that most of the proteins that are synthesized in the eukaryotic cell may fold sequentially. This proposed folding mechanism in vivo is particularly advantageous to the organism. In the absence of chaperones, the probability that a sequentially folding protein will misfold is reduced significantly. The problem we address here is devising a procedure that would differentiate between the two types of folding patterns. Footprints of sequential folding may be found in structures where consecutive fragments of the chain interact with each other. In such cases, the folding complexity may be viewed as being lower. On the other hand, higher folding complexity suggests that at least a portion of the polypeptide backbone folds back upon itself to form three-dimensional (3D) interactions with noncontiguous portion(s) of the chain. Hence, we look at the mechanism of folding of the molecule via analysis of its complexity, that is, through the 3D interactions formed by contiguous segments on the polypeptide chain. To computationally splice the structure into consecutively interacting fragments, we either cut it into compact hydrophobic folding units or into a set of hypothetical, transient, highly populated, contiguous fragments ("building blocks" of the structure). In sequential folding, successive building blocks interact with each other from the amino to the carboxy terminus of the polypeptide chain. Consequently, the results of the parsing differentiate between sequentially vs. nonsequentially folded chains. The automated assessment of the folding complexity provides insight into both the likelihood of misfolding and the kinetic folding rate of the given protein. In terms of the funnel free energy landscape theory, a protein that truly follows the mechanism of sequential folding, in principle, encounters smoother free energy barriers

  6. Guiding the folding pathway of DNA origami

    NASA Astrophysics Data System (ADS)

    Dunn, Katherine E.; Dannenberg, Frits; Ouldridge, Thomas E.; Kwiatkowska, Marta; Turberfield, Andrew J.; Bath, Jonathan

    2015-09-01

    DNA origami is a robust assembly technique that folds a single-stranded DNA template into a target structure by annealing it with hundreds of short `staple' strands. Its guiding design principle is that the target structure is the single most stable configuration. The folding transition is cooperative and, as in the case of proteins, is governed by information encoded in the polymer sequence. A typical origami folds primarily into the desired shape, but misfolded structures can kinetically trap the system and reduce the yield. Although adjusting assembly conditions or following empirical design rules can improve yield, well-folded origami often need to be separated from misfolded structures. The problem could in principle be avoided if assembly pathway and kinetics were fully understood and then rationally optimized. To this end, here we present a DNA origami system with the unusual property of being able to form a small set of distinguishable and well-folded shapes that represent discrete and approximately degenerate energy minima in a vast folding landscape, thus allowing us to probe the assembly process. The obtained high yield of well-folded origami structures confirms the existence of efficient folding pathways, while the shape distribution provides information about individual trajectories through the folding landscape. We find that, similarly to protein folding, the assembly of DNA origami is highly cooperative; that reversible bond formation is important in recovering from transient misfoldings; and that the early formation of long-range connections can very effectively enforce particular folds. We use these insights to inform the design of the system so as to steer assembly towards desired structures. Expanding the rational design process to include the assembly pathway should thus enable more reproducible synthesis, particularly when targeting more complex structures. We anticipate that this expansion will be essential if DNA origami is to continue its

  7. The unsolved chapter of vocal fold scars and how tissue engineering could help us solve the problem.

    PubMed

    Graupp, M; Bachna-Rotter, S; Gerstenberger, C; Friedrich, G; Fröhlich-Sorger, E; Kiesler, K; Gugatschka, M

    2016-09-01

    Vocal fold scarring is a relatively small field in scar research with prerequisites found nowhere else. The deterioration of the delicate tri-layered micro-structure of the epithelium of the vocal folds leads to impaired vibration characteristics resulting in a permanent hoarse and breathy voice. Tissue engineering approaches could help to restore the pre-injury status. Despite a considerable progress in this field during the last years, routine clinical applications are not available so far. One reason might be that vocal fold fibroblasts, as the responsible cell type for fibrogenesis, have very particular properties that are only poorly characterized. Moreover, in vivo trials are costly and time consuming and a representative in vitro model does not exist so far. These particular circumstances lead to innovative in vitro strategies and concepts such as macro-molecular crowding that can also be applied in adjacent fields.

  8. Nonlinear Models for Protein Folding and Function

    NASA Astrophysics Data System (ADS)

    Cruzeiro, L.

    Earlier a specific kinetic process for reproducible protein folding was proposed according to which the nascent chain is helical and the first step in in vivo protein folding is the bending of the initial helix at specific amino acid sites. Here the theoretical feasibility of this kinetic process is tested. To that end, two proteins, one belonging to the mainly α class and the other belonging to the α/β class, are selected and targeted molecular dynamics is applied to generate folding pathways for those two proteins, starting from two well defined initial conformations: a fully extended and a α-helical conformation. Not only are the native states closer to an initial helical structure for both proteins but also the pathways from the α-helical initial conformation to the native state have lower potential energy than the pathways that start from the fully extended conformation. For the α/β protein, 30% (40%) of the pathways from an initial α-helix (fully extended) structure lead to unentangled native folds, a success rate that can be increased to 85% by the introduction of a putative intermediate structure. These results lend support to the kinetic process proposed and open up a new direction in which to look for a solution to the protein folding problem. The chapter ends with a section that emphasizes the formal similarities between the dynamics quantum vibrational excited states in proteins and electrons in nonlinear lattices.

  9. Progress in fold recognition.

    PubMed

    Flöckner, H; Braxenthaler, M; Lackner, P; Jaritz, M; Ortner, M; Sippl, M J

    1995-11-01

    The prediction experiment reveals that fold recognition has become a powerful tool in structural biology. We applied our fold recognition technique to 13 target sequences. In two cases, replication terminating protein and prosequence of subtilisin, the predicted structures are very similar to the experimentally determined folds. For the first time, in a public blind test, the unknown structures of proteins have been predicted ahead of experiment to an accuracy approaching molecular detail. In two other cases the approximate folds have been predicted correctly. According to the assessors there were 12 recognizable folds among the target proteins. In our postprediction analysis we find that in 7 cases our fold recognition technique is successful. In several of the remaining cases the predicted folds have interesting features in common with the experimental results. We present our procedure, discuss the results, and comment on several fundamental and technical problems encountered in fold recognition.

  10. Folding Beauties

    ERIC Educational Resources Information Center

    Berman, Leah Wrenn

    2006-01-01

    This article has its genesis in an MAA mini-course on origami, where a way to get a parabola by folding paper was presented. This article discusses the methods and mathematics of other curves obtained by paper-folding.

  11. THE SOLAR ABUNDANCE PROBLEM: THE EFFECT OF THE TURBULENT KINETIC FLUX ON THE SOLAR ENVELOPE MODEL

    SciTech Connect

    Zhang, Q. S.

    2014-06-01

    Recent three-dimensional (3D) simulations have shown that the turbulent kinetic flux (TKF) is significant. We discuss the effects of TKF on the size of the convection zone and find that the TKF may help solve the solar abundance problem. The solar abundance problem is that, with new abundances, the solar convection zone depth, the sound speed in the radiative interior, the helium abundance, and the density in the convective envelope are not in agreement with helioseismic inversions. We have performed Monte Carlo simulations on solar convective envelope models with different profiles of TKF to test its effects. The solar abundance problem is revealed in the standard solar convective envelope model with AGSS09 composition, which shows significant differences (∼10)) in density from the helioseismic inversions, but the differences in the model with the old composition GN93 is small (∼0.5)). In the testing models with a different TKF imposed, it is found that the density profile is sensitive to the value of TKF at the base of the convective envelope and insensitive to the structure of TKF in the convection zone. The required value of turbulent kinetic luminosity at the base is about –13% to – 19% L {sub ☉}. Comparing with the 3D simulations, this value is plausible. This study is for the solar convective envelope only. Evolutionary solar models with TKF are required to investigat the effects of TKF on the solar interior structure below the convection zone and the whole solar abundance problem, but the profile of the TKF in the overshoot region is necessary.

  12. LSENS: A General Chemical Kinetics and Sensitivity Analysis Code for homogeneous gas-phase reactions. Part 3: Illustrative test problems

    NASA Technical Reports Server (NTRS)

    Bittker, David A.; Radhakrishnan, Krishnan

    1994-01-01

    LSENS, the Lewis General Chemical Kinetics and Sensitivity Analysis Code, has been developed for solving complex, homogeneous, gas-phase chemical kinetics problems and contains sensitivity analysis for a variety of problems, including nonisothermal situations. This report is part 3 of a series of three reference publications that describe LSENS, provide a detailed guide to its usage, and present many example problems. Part 3 explains the kinetics and kinetics-plus-sensitivity analysis problems supplied with LSENS and presents sample results. These problems illustrate the various capabilities of, and reaction models that can be solved by, the code and may provide a convenient starting point for the user to construct the problem data file required to execute LSENS. LSENS is a flexible, convenient, accurate, and efficient solver for chemical reaction problems such as static system; steady, one-dimensional, inviscid flow; reaction behind incident shock wave, including boundary layer correction; and perfectly stirred (highly backmixed) reactor. In addition, the chemical equilibrium state can be computed for the following assigned states: temperature and pressure, enthalpy and pressure, temperature and volume, and internal energy and volume. For static problems the code computes the sensitivity coefficients of the dependent variables and their temporal derivatives with respect to the initial values of the dependent variables and/or the three rate coefficient parameters of the chemical reactions.

  13. Early Events in RNA Folding

    NASA Astrophysics Data System (ADS)

    Thirumalai, D.; Lee, Namkyung; Woodson, Sarah A.; Klimov, Dk

    2001-10-01

    We describe a conceptual framework for understanding the way large RNA molecules fold based on the notion that their free-energy landscape is rugged. A key prediction of our theory is that RNA folding can be described by the kinetic partitioning mechanism (KPM). According to KPM a small fraction of molecules folds rapidly to the native state whereas the remaining fraction is kinetically trapped in a low free-energy non-native state. This model provides a unified description of the way RNA and proteins fold. Single-molecule experiments on Tetrahymena ribozyme, which directly validate our theory, are analyzed using KPM. We also describe the earliest events that occur on microsecond time scales in RNA folding. These must involve collapse of RNA molecules that are mediated by counterion-condensation. Estimates of time scales for the initial events in RNA folding are provided for the Tetrahymena ribozyme.

  14. Determination of three-dimensional structures of proteins from interproton distance data by dynamical simulated annealing from a random array of atoms. Circumventing problems associated with folding.

    PubMed

    Nilges, M; Clore, G M; Gronenborn, A M

    1988-10-24

    A new real space method, based on the principles of simulated annealing, is presented for determining protein structures on the basis of interproton distance restraints derived from NMR data. The method circumvents the folding problem associated with all real space methods described to date, by starting from a completely random array of atoms and introducing the force constants for the covalent, interproton distance and repulsive van der Waals terms in the target function appropriately. The system is simulated at high temperature by solving Newton's equations of motion. As the values of all force constants are very low during the early stages of the simulation, energy barriers between different folds of the protein can be overcome, and the global minimum of the target function is reliably located. Further, because the atoms are initially only weakly coupled, they can move essentially independently to satisfy the restraints. The method is illustrated using two examples of small proteins, namely crambin (46 residues) and potato carboxypeptidase inhibitor (39 residues).

  15. A hybrid computer program for rapidly solving flowing or static chemical kinetic problems involving many chemical species

    NASA Technical Reports Server (NTRS)

    Mclain, A. G.; Rao, C. S. R.

    1976-01-01

    A hybrid chemical kinetic computer program was assembled which provides a rapid solution to problems involving flowing or static, chemically reacting, gas mixtures. The computer program uses existing subroutines for problem setup, initialization, and preliminary calculations and incorporates a stiff ordinary differential equation solution technique. A number of check cases were recomputed with the hybrid program and the results were almost identical to those previously obtained. The computational time saving was demonstrated with a propane-oxygen-argon shock tube combustion problem involving 31 chemical species and 64 reactions. Information is presented to enable potential users to prepare an input data deck for the calculation of a problem.

  16. Some current problems in perovskite nano-ferroelectrics and multiferroics: kinetically-limited systems of finite lateral size

    NASA Astrophysics Data System (ADS)

    Scott, James F.; Schilling, Alina; Rowley, S. E.; Gregg, J. Marty

    2015-06-01

    We describe some unsolved problems of current interest; these involve quantum critical points in ferroelectrics and problems which are not amenable to the usual density functional theory, nor to classical Landau free energy approaches (they are kinetically limited), nor even to the Landau-Kittel relationship for domain size (they do not satisfy the assumption of infinite lateral diameter) because they are dominated by finite aperiodic boundary conditions.

  17. An instrument for fast acquisition of fluorescence decay curves at picosecond resolution designed for ``double kinetics'' experiments: Application to fluorescence resonance excitation energy transfer study of protein folding

    NASA Astrophysics Data System (ADS)

    Ishay, Eldad Ben; Hazan, Gershon; Rahamim, Gil; Amir, Dan; Haas, Elisha

    2012-08-01

    The information obtained by studying fluorescence decay of labeled biopolymers is a major resource for understanding the dynamics of their conformations and interactions. The lifetime of the excited states of probes attached to macromolecules is in the nanosecond time regime, and hence, a series of snapshot decay curves of such probes might - in principle - yield details of fast changes of ensembles of labeled molecules down to sub-microsecond time resolution. Hence, a major current challenge is the development of instruments for the low noise detection of fluorescence decay curves within the shortest possible time intervals. Here, we report the development of an instrument, picosecond double kinetics apparatus, that enables recording of multiple fluorescence decay curves with picosecond excitation pulses over wide spectral range during microsecond data collection for each curve. The design is based on recording and averaging multiphoton pulses of fluorescence decay using a fast 13 GHz oscilloscope during microsecond time intervals at selected time points over the course of a chemical reaction or conformational transition. We tested this instrument in a double kinetics experiment using reference probes (N-acetyl-tryptophanamide). Very low stochastic noise level was attained, and reliable multi-parameter analysis such as derivation of distance distributions from time resolved FRET (fluorescence resonance excitation energy transfer) measurements was achieved. The advantage of the pulse recording and averaging approach used here relative to double kinetics methods based on the established time correlated single photon counting method, is that in the pulse recording approach, averaging of substantially fewer kinetic experiments is sufficient for obtaining the data. This results in a major reduction in the consumption of labeled samples, which in many cases, enables the performance of important experiments that were not previously feasible.

  18. Solvent–amino acid interaction energies in three-dimensional-lattice Monte Carlo simulations of a model 27-mer protein: Folding thermodynamics and kinetics

    PubMed Central

    Leonhard, Kai; Prausnitz, John M.; Radke, Clayton J.

    2004-01-01

    Amino acid residue–solvent interactions are required for lattice Monte Carlo simulations of model proteins in water. In this study, we propose an interaction-energy scale that is based on the interaction scale by Miyazawa and Jernigan. It permits systematic variation of the amino acid–solvent interactions by introducing a contrast parameter for the hydrophobicity, Cs, and a mean attraction parameter for the amino acids, ω. Changes in the interaction energies strongly affect many protein properties. We present an optimized energy parameter set for best representing realistic behavior typical for many proteins (fast folding and high cooperativity for single chains). Our optimal parameters feature a much weaker hydrophobicity contrast and mean attraction than does the original interaction scale. The proposed interaction scale is designed for calculating the behavior of proteins in bulk and at interfaces as a function of solvent characteristics, as well as protein size and sequence. PMID:14739322

  19. Evolutionary optimization of protein folding.

    PubMed

    Debès, Cédric; Wang, Minglei; Caetano-Anollés, Gustavo; Gräter, Frauke

    2013-01-01

    Nature has shaped the make up of proteins since their appearance, [Formula: see text]3.8 billion years ago. However, the fundamental drivers of structural change responsible for the extraordinary diversity of proteins have yet to be elucidated. Here we explore if protein evolution affects folding speed. We estimated folding times for the present-day catalog of protein domains directly from their size-modified contact order. These values were mapped onto an evolutionary timeline of domain appearance derived from a phylogenomic analysis of protein domains in 989 fully-sequenced genomes. Our results show a clear overall increase of folding speed during evolution, with known ultra-fast downhill folders appearing rather late in the timeline. Remarkably, folding optimization depends on secondary structure. While alpha-folds showed a tendency to fold faster throughout evolution, beta-folds exhibited a trend of folding time increase during the last [Formula: see text]1.5 billion years that began during the "big bang" of domain combinations. As a consequence, these domain structures are on average slow folders today. Our results suggest that fast and efficient folding of domains shaped the universe of protein structure. This finding supports the hypothesis that optimization of the kinetic and thermodynamic accessibility of the native fold reduces protein aggregation propensities that hamper cellular functions. PMID:23341762

  20. Criteria for folding in structure-based models of proteins

    NASA Astrophysics Data System (ADS)

    Wołek, Karol; Cieplak, Marek

    2016-05-01

    In structure-based models of proteins, one often assumes that folding is accomplished when all contacts are established. This assumption may frequently lead to a conceptual problem that folding takes place in a temperature region of very low thermodynamic stability, especially when the contact map used is too sparse. We consider six different structure-based models and show that allowing for a small, but model-dependent, percentage of the native contacts not being established boosts the folding temperature substantially while affecting the time scales of folding only in a minor way. We also compare other properties of the six models. We show that the choice of the description of the backbone stiffness has a substantial effect on the values of characteristic temperatures that relate both to equilibrium and kinetic properties. Models without any backbone stiffness (like the self-organized polymer) are found to perform similar to those with the stiffness, including in the studies of stretching.

  1. Moment-Based Accelerators for Kinetic Problems with Application to Inertial Confinement Fusion

    NASA Astrophysics Data System (ADS)

    Taitano, William Tsubasa-Tsutsui

    In inertial confinement fusion (ICF), the kinetic ion and charge separation field effects may play a significant role in the difference between the measured neutron yield in experiments and the predicted yield from fluid codes. Two distinct of approaches exists in modeling plasma physics phenomena: fluid and kinetic approaches. While the fluid approach is computationally less expensive, robust closures are difficult to obtain for a wide separation in temperature and density. While the kinetic approach is a closed system, it resolves the full 6D phase space and classic explicit numerical schemes restrict both the spatial and time-step size to a point where the method becomes intractable. Classic implicit system require the storage and inversion of a very large linear system which also becomes intractable. This dissertation will develop a new implicit method based on an emerging moment-based accelerator which allows one to step over stiff kinetic time-scales. The new method converges the solution per time-step stably and efficiently compared to a standard Picard iteration. This new algorithm will be used to investigate mixing in Omega ICF fuel-pusher interface at early time of the implosion process, fully kinetically.

  2. PREFACE Protein folding: lessons learned and new frontiers Protein folding: lessons learned and new frontiers

    NASA Astrophysics Data System (ADS)

    Pappu, Rohit V.; Nussinov, Ruth

    2009-03-01

    In appropriate physiological milieux proteins spontaneously fold into their functional three-dimensional structures. The amino acid sequences of functional proteins contain all the information necessary to specify the folds. This remarkable observation has spawned research aimed at answering two major questions. (1) Of all the conceivable structures that a protein can adopt, why is the ensemble of native-like structures the most favorable? (2) What are the paths by which proteins manage to robustly and reproducibly fold into their native structures? Anfinsen's thermodynamic hypothesis has guided the pursuit of answers to the first question whereas Levinthal's paradox has influenced the development of models for protein folding dynamics. Decades of work have led to significant advances in the folding problem. Mean-field models have been developed to capture our current, coarse grain understanding of the driving forces for protein folding. These models are being used to predict three-dimensional protein structures from sequence and stability profiles as a function of thermodynamic and chemical perturbations. Impressive strides have also been made in the field of protein design, also known as the inverse folding problem, thereby testing our understanding of the determinants of the fold specificities of different sequences. Early work on protein folding pathways focused on the specific sequence of events that could lead to a simplification of the search process. However, unifying principles proved to be elusive. Proteins that show reversible two-state folding-unfolding transitions turned out to be a gift of natural selection. Focusing on these simple systems helped researchers to uncover general principles regarding the origins of cooperativity in protein folding thermodynamics and kinetics. On the theoretical front, concepts borrowed from polymer physics and the physics of spin glasses led to the development of a framework based on energy landscape theories. These

  3. Suitable combination of promoter and micellar catalyst for kilo fold rate acceleration on benzaldehyde to benzoic acid conversion in aqueous media at room temperature: A kinetic approach

    NASA Astrophysics Data System (ADS)

    Ghosh, Aniruddha; Saha, Rumpa; Ghosh, Sumanta K.; Mukherjee, Kakali; Saha, Bidyut

    2013-05-01

    The kinetics of oxidation of benzaldehyde by chromic acid in aqueous and aqueous surfactant (sodium dodecyl sulfate, SDS, alkyl phenyl polyethylene glycol, Triton X-100 and N-cetylpyridinium chloride, CPC) media have been investigated in the presence of promoter at 303 K. The pseudo-first-order rate constants (kobs) were determined from a logarithmic plot of absorbance as a function time. The rate constants were found to increase with introduction of heteroaromatic nitrogen base promoters such as Picolinic acid (PA), 2,2'-bipyridine (bipy) and 1,10-phenanthroline (phen). The product benzoic acid has been characterized by conventional melting point experiment, NMR, HRMS and FTIR spectral analysis. The mechanism of both unpromoted and promoted reaction path has been proposed for the reaction. In presence of the anionic surfactant SDS, cationic surfactant CPC and neutral surfactant TX-100 the reaction can undergo simultaneously in both aqueous and micellar phase with an enhanced rate of oxidation in the micellar phase. Both SDS and TX-100 produce normal micellar effect whereas CPC produce reverse micellar effect in the presence of benzaldehyde. The observed net enhancement of rate effects has been explained by considering the hydrophobic and electrostatic interaction between the surfactants and reactants. SDS and bipy combination is the suitable one for benzaldehyde oxidation.

  4. Suitable combination of promoter and micellar catalyst for kilo fold rate acceleration on benzaldehyde to benzoic acid conversion in aqueous media at room temperature: a kinetic approach.

    PubMed

    Ghosh, Aniruddha; Saha, Rumpa; Ghosh, Sumanta K; Mukherjee, Kakali; Saha, Bidyut

    2013-05-15

    The kinetics of oxidation of benzaldehyde by chromic acid in aqueous and aqueous surfactant (sodium dodecyl sulfate, SDS, alkyl phenyl polyethylene glycol, Triton X-100 and N-cetylpyridinium chloride, CPC) media have been investigated in the presence of promoter at 303 K. The pseudo-first-order rate constants (kobs) were determined from a logarithmic plot of absorbance as a function time. The rate constants were found to increase with introduction of heteroaromatic nitrogen base promoters such as Picolinic acid (PA), 2,2'-bipyridine (bipy) and 1,10-phenanthroline (phen). The product benzoic acid has been characterized by conventional melting point experiment, NMR, HRMS and FTIR spectral analysis. The mechanism of both unpromoted and promoted reaction path has been proposed for the reaction. In presence of the anionic surfactant SDS, cationic surfactant CPC and neutral surfactant TX-100 the reaction can undergo simultaneously in both aqueous and micellar phase with an enhanced rate of oxidation in the micellar phase. Both SDS and TX-100 produce normal micellar effect whereas CPC produce reverse micellar effect in the presence of benzaldehyde. The observed net enhancement of rate effects has been explained by considering the hydrophobic and electrostatic interaction between the surfactants and reactants. SDS and bipy combination is the suitable one for benzaldehyde oxidation.

  5. Peptide Folding: Many Routes to the Native State.

    NASA Astrophysics Data System (ADS)

    Roitberg, Adrian; Simmerling, Carlos

    2002-03-01

    Protein folding is only one example of an extremely complex kinetic process that is poorly understood due to experimental limitations. Several groups have published simulations of early folding stages in atomic detail, but kinetic processes are probabilistic in nature and single simulations may have little value. Clusters of computers can rapidly provide multiple data sets that can be combined to obtain greater insight. We have carried out the computational analog of temperature-jump protein folding. An ensemble of structures that represent the unfolded state is generated at 800K by molecular dynamics. Each structure was distributed to an individual node in the cluster, quenched to 298K and monitored during MD simulation. We have accumulated 2.3 microseconds of folding time in this way. While it is possible to perform this procedure using a single computer, perfect scaling is obtained with the cluster and the problem is solved far more quickly. We carried out a preliminary study on a nonapeptide fragment of influenza virus hemagluttinin using a Generalized Born aqueous solvation model. Under these conditions, the peptide has a high tendency to form a well-defined structure stabilized predominantly by backbone hydrogen bonds with a structure similar to that observed experimentally in the intact protein. Folding times of the ensemble of 188 unfolded structures varied from 2ps to over 100ns and at least three exponential curves are required to fit this data. Analysis of trajectories revealed four distinct folding pathways with kinetic properties that fit these individual curves, with two different pathways contributing to the slowest folding events. Folding time varies by an order of magnitude even on the same pathway. We obtained the important and potentially general result that folding kinetics depends strongly on the details of preparation of this ensemble. To the best of our knowledge, this is the first time such detailed analysis of protein folding landscapes has

  6. Quantifying Nonnative Interactions in the Protein-Folding Free-Energy Landscape.

    PubMed

    Mouro, Paulo Ricardo; de Godoi Contessoto, Vinícius; Chahine, Jorge; Junio de Oliveira, Ronaldo; Pereira Leite, Vitor Barbanti

    2016-07-26

    Protein folding is a central problem in biological physics. Energetic roughness is an important aspect that controls protein-folding stability and kinetics. The roughness is associated with conflicting interactions in the protein and is also known as frustration. Recent studies indicate that an addition of a small amount of energetic frustration may enhance folding speed for certain proteins. In this study, we have investigated the conditions under which frustration increases the folding rate. We used a Cα structure-based model to simulate a group of proteins. We found that the free-energy barrier at the transition state (ΔF) correlates with nonnative-contact variation (ΔA), and the simulated proteins are clustered according to their fold motifs. These findings are corroborated by the Clementi-Plotkin analytical model. As a consequence, the optimum frustration regime for protein folding can be predicted analytically. PMID:27463131

  7. Pseudoknots in RNA folding landscapes

    PubMed Central

    Kucharík, Marcel; Hofacker, Ivo L.; Stadler, Peter F.; Qin, Jing

    2016-01-01

    Motivation: The function of an RNA molecule is not only linked to its native structure, which is usually taken to be the ground state of its folding landscape, but also in many cases crucially depends on the details of the folding pathways such as stable folding intermediates or the timing of the folding process itself. To model and understand these processes, it is necessary to go beyond ground state structures. The study of rugged RNA folding landscapes holds the key to answer these questions. Efficient coarse-graining methods are required to reduce the intractably vast energy landscapes into condensed representations such as barrier trees or basin hopping graphs (BHG) that convey an approximate but comprehensive picture of the folding kinetics. So far, exact and heuristic coarse-graining methods have been mostly restricted to the pseudoknot-free secondary structures. Pseudoknots, which are common motifs and have been repeatedly hypothesized to play an important role in guiding folding trajectories, were usually excluded. Results: We generalize the BHG framework to include pseudoknotted RNA structures and systematically study the differences in predicted folding behavior depending on whether pseudoknotted structures are allowed to occur as folding intermediates or not. We observe that RNAs with pseudoknotted ground state structures tend to have more pseudoknotted folding intermediates than RNAs with pseudoknot-free ground state structures. The occurrence and influence of pseudoknotted intermediates on the folding pathway, however, appear to depend very strongly on the individual RNAs so that no general rule can be inferred. Availability and implementation: The algorithms described here are implemented in C++ as standalone programs. Its source code and Supplemental material can be freely downloaded from http://www.tbi.univie.ac.at/bhg.html. Contact: qin@bioinf.uni-leipzig.de Supplementary information: Supplementary data are available at Bioinformatics online. PMID

  8. Atomic-level description of ubiquitin folding

    PubMed Central

    Piana, Stefano; Lindorff-Larsen, Kresten; Shaw, David E.

    2013-01-01

    Equilibrium molecular dynamics simulations, in which proteins spontaneously and repeatedly fold and unfold, have recently been used to help elucidate the mechanistic principles that underlie the folding of fast-folding proteins. The extent to which the conclusions drawn from the analysis of such proteins, which fold on the microsecond timescale, apply to the millisecond or slower folding of naturally occurring proteins is, however, unclear. As a first attempt to address this outstanding issue, we examine here the folding of ubiquitin, a 76-residue-long protein found in all eukaryotes that is known experimentally to fold on a millisecond timescale. Ubiquitin folding has been the subject of many experimental studies, but its slow folding rate has made it difficult to observe and characterize the folding process through all-atom molecular dynamics simulations. Here we determine the mechanism, thermodynamics, and kinetics of ubiquitin folding through equilibrium atomistic simulations. The picture emerging from the simulations is in agreement with a view of ubiquitin folding suggested from previous experiments. Our findings related to the folding of ubiquitin are also consistent, for the most part, with the folding principles derived from the simulation of fast-folding proteins, suggesting that these principles may be applicable to a wider range of proteins. PMID:23503848

  9. Reduced alphabet for protein folding prediction.

    PubMed

    Huang, Jitao T; Wang, Titi; Huang, Shanran R; Li, Xin

    2015-04-01

    What are the key building blocks that would have been needed to construct complex protein folds? This is an important issue for understanding protein folding mechanism and guiding de novo protein design. Twenty naturally occurring amino acids and eight secondary structures consist of a 28-letter alphabet to determine folding kinetics and mechanism. Here we predict folding kinetic rates of proteins from many reduced alphabets. We find that a reduced alphabet of 10 letters achieves good correlation with folding rates, close to the one achieved by full 28-letter alphabet. Many other reduced alphabets are not significantly correlated to folding rates. The finding suggests that not all amino acids and secondary structures are equally important for protein folding. The foldable sequence of a protein could be designed using at least 10 folding units, which can either promote or inhibit protein folding. Reducing alphabet cardinality without losing key folding kinetic information opens the door to potentially faster machine learning and data mining applications in protein structure prediction, sequence alignment and protein design.

  10. Application of a four-step HMX kinetic model to an impact-induced fraction ignition problems

    SciTech Connect

    Perry, William L; Gunderson, Jake A; Dickson, Peter M

    2010-01-01

    There has been a long history of interest in the decomposition kinetics of HMX and HMX-based formulations due to the widespread use of this explosive in high performance systems. The kinetics allow us to predict, or attempt to predict, the behavior of the explosive when subjected to thermal hazard scenarios that lead to ignition via impact, spark, friction or external heat. The latter, commonly referred to as 'cook off', has been widely studied and contemporary kinetic and transport models accurately predict time and location of ignition for simple geometries. However, there has been relatively little attention given to the problem of localized ignition that results from the first three ignition sources of impact, spark and friction. The use of a zero-order single-rate expression describing the exothermic decomposition of explosives dates to the early work of Frank-Kamanetskii in the late 1930s and continued through the 60's and 70's. This expression provides very general qualitative insight, but cannot provide accurate spatial or timing details of slow cook off ignition. In the 70s, Catalano, et al., noted that single step kinetics would not accurately predict time to ignition in the one-dimensional time to explosion apparatus (ODTX). In the early 80s, Tarver and McGuire published their well-known three step kinetic expression that included an endothermic decomposition step. This scheme significantly improved the accuracy of ignition time prediction for the ODTX. However, the Tarver/McGuire model could not produce the internal temperature profiles observed in the small-scale radial experiments nor could it accurately predict the location of ignition. Those factors are suspected to significantly affect the post-ignition behavior and better models were needed. Brill, et al. noted that the enthalpy change due to the beta-delta crystal phase transition was similar to the assumed endothermic decomposition step in the Tarver/McGuire model. Henson, et al., deduced the

  11. Unfolded protein ensembles, folding trajectories, and refolding rate prediction.

    PubMed

    Das, A; Sin, B K; Mohazab, A R; Plotkin, S S

    2013-09-28

    Computer simulations can provide critical information on the unfolded ensemble of proteins under physiological conditions, by explicitly characterizing the geometrical properties of the diverse conformations that are sampled in the unfolded state. A general computational analysis across many proteins has not been implemented however. Here, we develop a method for generating a diverse conformational ensemble, to characterize properties of the unfolded states of intrinsically disordered or intrinsically folded proteins. The method allows unfolded proteins to retain disulfide bonds. We examined physical properties of the unfolded ensembles of several proteins, including chemical shifts, clustering properties, and scaling exponents for the radius of gyration with polymer length. A problem relating simulated and experimental residual dipolar couplings is discussed. We apply our generated ensembles to the problem of folding kinetics, by examining whether the ensembles of some proteins are closer geometrically to their folded structures than others. We find that for a randomly selected dataset of 15 non-homologous 2- and 3-state proteins, quantities such as the average root mean squared deviation between the folded structure and unfolded ensemble correlate with folding rates as strongly as absolute contact order. We introduce a new order parameter that measures the distance travelled per residue, which naturally partitions into a smooth "laminar" and subsequent "turbulent" part of the trajectory. This latter conceptually simple measure with no fitting parameters predicts folding rates in 0 M denaturant with remarkable accuracy (r = -0.95, p = 1 × 10(-7)). The high correlation between folding times and sterically modulated, reconfigurational motion supports the rapid collapse of proteins prior to the transition state as a generic feature in the folding of both two-state and multi-state proteins. This method for generating unfolded ensembles provides a powerful approach to

  12. Unfolded protein ensembles, folding trajectories, and refolding rate prediction

    NASA Astrophysics Data System (ADS)

    Das, A.; Sin, B. K.; Mohazab, A. R.; Plotkin, S. S.

    2013-09-01

    Computer simulations can provide critical information on the unfolded ensemble of proteins under physiological conditions, by explicitly characterizing the geometrical properties of the diverse conformations that are sampled in the unfolded state. A general computational analysis across many proteins has not been implemented however. Here, we develop a method for generating a diverse conformational ensemble, to characterize properties of the unfolded states of intrinsically disordered or intrinsically folded proteins. The method allows unfolded proteins to retain disulfide bonds. We examined physical properties of the unfolded ensembles of several proteins, including chemical shifts, clustering properties, and scaling exponents for the radius of gyration with polymer length. A problem relating simulated and experimental residual dipolar couplings is discussed. We apply our generated ensembles to the problem of folding kinetics, by examining whether the ensembles of some proteins are closer geometrically to their folded structures than others. We find that for a randomly selected dataset of 15 non-homologous 2- and 3-state proteins, quantities such as the average root mean squared deviation between the folded structure and unfolded ensemble correlate with folding rates as strongly as absolute contact order. We introduce a new order parameter that measures the distance travelled per residue, which naturally partitions into a smooth "laminar" and subsequent "turbulent" part of the trajectory. This latter conceptually simple measure with no fitting parameters predicts folding rates in 0 M denaturant with remarkable accuracy (r = -0.95, p = 1 × 10-7). The high correlation between folding times and sterically modulated, reconfigurational motion supports the rapid collapse of proteins prior to the transition state as a generic feature in the folding of both two-state and multi-state proteins. This method for generating unfolded ensembles provides a powerful approach to

  13. In-Space Propulsion, Logistics Reduction, and Evaluation of Steam Reformer Kinetics: Problems and Prospects

    NASA Technical Reports Server (NTRS)

    Jaworske, D. A.; Palaszewski, B. A.; Kulis, M. J.; Gokoglu, S. A.

    2015-01-01

    Human space missions generate waste materials. A 70-kg crewmember creates a waste stream of 1 kg per day, and a four-person crew on a deep space habitat for a 400+ day mission would create over 1600 kg of waste. Converted into methane, the carbon could be used as a fuel for propulsion or power. The NASA Advanced Exploration Systems (AES) Logistics Reduction and Repurposing (LRR) project is investing in space resource utilization with an emphasis on repurposing logistics materials for useful purposes and has selected steam reforming among many different competitive processes as the preferred method for repurposing organic waste into methane. Already demonstrated at the relevant processing rate of 5.4 kg of waste per day, high temperature oxygenated steam consumes waste and produces carbon dioxide, carbon monoxide, and hydrogen which can then be converted into methane catalytically. However, the steam reforming process has not been studied in microgravity. Data are critically needed to understand the mechanisms that allow use of steam reforming in a reduced gravity environment. This paper reviews the relevant literature, identifies gravity-dependent mechanisms within the steam gasification process, and describes an innovative experiment to acquire the crucial kinetic information in a small-scale reactor specifically designed to operate within the requirements of a reduced gravity aircraft flight. The experiment will determine if the steam reformer process is mass-transport limited, and if so, what level of forced convection will be needed to obtain performance comparable to that in 1-g.

  14. Improving protein fold recognition by random forest

    PubMed Central

    2014-01-01

    Background Recognizing the correct structural fold among known template protein structures for a target protein (i.e. fold recognition) is essential for template-based protein structure modeling. Since the fold recognition problem can be defined as a binary classification problem of predicting whether or not the unknown fold of a target protein is similar to an already known template protein structure in a library, machine learning methods have been effectively applied to tackle this problem. In our work, we developed RF-Fold that uses random forest - one of the most powerful and scalable machine learning classification methods - to recognize protein folds. Results RF-Fold consists of hundreds of decision trees that can be trained efficiently on very large datasets to make accurate predictions on a highly imbalanced dataset. We evaluated RF-Fold on the standard Lindahl's benchmark dataset comprised of 976 × 975 target-template protein pairs through cross-validation. Compared with 17 different fold recognition methods, the performance of RF-Fold is generally comparable to the best performance in fold recognition of different difficulty ranging from the easiest family level, the medium-hard superfamily level, and to the hardest fold level. Based on the top-one template protein ranked by RF-Fold, the correct recognition rate is 84.5%, 63.4%, and 40.8% at family, superfamily, and fold levels, respectively. Based on the top-five template protein folds ranked by RF-Fold, the correct recognition rate increases to 91.5%, 79.3% and 58.3% at family, superfamily, and fold levels. Conclusions The good performance achieved by the RF-Fold demonstrates the random forest's effectiveness for protein fold recognition. PMID:25350499

  15. Learning To Fold Proteins Using Energy Landscape Theory

    PubMed Central

    Schafer, N.P.; Kim, B.L.; Zheng, W.; Wolynes, P.G.

    2014-01-01

    This review is a tutorial for scientists interested in the problem of protein structure prediction, particularly those interested in using coarse-grained molecular dynamics models that are optimized using lessons learned from the energy landscape theory of protein folding. We also present a review of the results of the AMH/AMC/AMW/AWSEM family of coarse-grained molecular dynamics protein folding models to illustrate the points covered in the first part of the article. Accurate coarse-grained structure prediction models can be used to investigate a wide range of conceptual and mechanistic issues outside of protein structure prediction; specifically, the paper concludes by reviewing how AWSEM has in recent years been able to elucidate questions related to the unusual kinetic behavior of artificially designed proteins, multidomain protein misfolding, and the initial stages of protein aggregation. PMID:25308991

  16. Let Them Fold

    ERIC Educational Resources Information Center

    Grant, Nicholas; Tobin, Alexander

    1972-01-01

    Directions are given for seven activities involving the folding of paper strips to illustrate geometric concepts. Properties of pentagons, triangles, hexagons, and Mobius bands resulting from the various foldings are discussed. (DT)

  17. Folding of viscous sheets and filaments

    NASA Astrophysics Data System (ADS)

    Skorobogatiy, M.; Mahadevan, L.

    2000-12-01

    We consider the nonlinear folding behavior of a viscous filament or a sheet under the influence of an external force such as gravity. Everyday examples of this phenomenon are provided by the periodic folding of a sheet of honey as it impinges on toast, or the folding of a stream of shampoo as it falls on one's hand. To understand the evolution of a fold, we formulate and solve a free-boundary problem for the phenomenon, give scaling laws for the size of the folds and the frequency with which they are laid out, and verify these experimentally.

  18. Integrating Kinetic Model of E. coli with Genome Scale Metabolic Fluxes Overcomes Its Open System Problem and Reveals Bistability in Central Metabolism

    PubMed Central

    Mannan, Ahmad A.; Toya, Yoshihiro; Shimizu, Kazuyuki; McFadden, Johnjoe; Kierzek, Andrzej M.; Rocco, Andrea

    2015-01-01

    An understanding of the dynamics of the metabolic profile of a bacterial cell is sought from a dynamical systems analysis of kinetic models. This modelling formalism relies on a deterministic mathematical description of enzyme kinetics and their metabolite regulation. However, it is severely impeded by the lack of available kinetic information, limiting the size of the system that can be modelled. Furthermore, the subsystem of the metabolic network whose dynamics can be modelled is faced with three problems: how to parameterize the model with mostly incomplete steady state data, how to close what is now an inherently open system, and how to account for the impact on growth. In this study we address these challenges of kinetic modelling by capitalizing on multi-‘omics’ steady state data and a genome-scale metabolic network model. We use these to generate parameters that integrate knowledge embedded in the genome-scale metabolic network model, into the most comprehensive kinetic model of the central carbon metabolism of E. coli realized to date. As an application, we performed a dynamical systems analysis of the resulting enriched model. This revealed bistability of the central carbon metabolism and thus its potential to express two distinct metabolic states. Furthermore, since our model-informing technique ensures both stable states are constrained by the same thermodynamically feasible steady state growth rate, the ensuing bistability represents a temporal coexistence of the two states, and by extension, reveals the emergence of a phenotypically heterogeneous population. PMID:26469081

  19. Foreland basins and fold belts

    SciTech Connect

    Macqueen, R.W.; Leckie, D.A. )

    1992-01-01

    The papers in this book describe six foreland basins and fold belts in terms of their regional setting, stratigraphy, tectonics, and structure, and their oil and gas systems. All of the basins show general similarities, but each differs significantly in detail from the others, posing something of a problem in terms of arriving at a 'typical' foreland basin and fold belt. Some are major hydrocarbon producers; others are not. The major characteristics of the six foreland basins and fold belts are summarized in Tables 1 through 5, which provide a convenient means of comparing and contrasting these basins and their hydrocarbon resources. The Western Canada foreland basin and fold belt serves as the type example for several reasons. These include: its setting and clear relationship to a major orogene of Mesozoic-Cenozoic age; the fact that it is uncomplicated by later overprinting, segmentation, or cover rocks unlike the Ouachita, Eastern Venezuela, and U.S. Rocky Mountain foreland basins and fold belts); the fact that there is a large volume of publicly available data on the basin and an active exploration and research community; and the fact that it has reasonable oil and gas reserves in a well-defined stratigraphic framework.

  20. STIS MAMA Fold Distribution

    NASA Astrophysics Data System (ADS)

    Wheeler, Thomas

    2012-10-01

    The performance of MAMA microchannel plates can be monitored using a MAMA fold distribution procedure. The fold distribution provides a measurement of the distribution of charge cloud sizes incident upon the anode giving some measure of change in the pulse-height distribution of the MCP and, therefore, MCP gain. This proposal executes the same steps as the STIS MAMA Fold Distribution, Proposal 12778, as Cycle 19.

  1. STIS MAMA Fold Distribution

    NASA Astrophysics Data System (ADS)

    Wheeler, Thomas

    2010-09-01

    The performance of MAMA microchannel plates can be monitored using a MAMA fold analysis procedure. The fold analysis provides a measurement of the distribution of charge cloud sizes incident upon the anode giving some measure of changes in the pulse-height distribution of the MCP and, therefore, MCP gain. This proposal executes the same steps as the STIS MAMA Fold Analysis {11863} during Cycle 17.

  2. STIS MAMA Fold Distribution

    NASA Astrophysics Data System (ADS)

    Wheeler, Thomas

    2009-07-01

    The performance of MAMA microchannel plates can be monitored using a MAMA fold analysis procedure. The fold analysis provides a measurement of the distribution of charge cloud sizes incident upon the anode giving some measure of changes in the pulse-height distribution of the MCP and, therefore, MCP gain. This proposal executes the same steps as the STIS MAMA Fold Analysis {10035} during Cycle 12.

  3. STIS MAMA Fold Distribution

    NASA Astrophysics Data System (ADS)

    Wheeler, Thomas

    2013-10-01

    The performance of MAMA microchannel plates can be monitored using a MAMA fold distribution procedure. The fold distribution provides a measurement of the distribution of charge cloud sizes incident upon the anode giving some measure of change in the pulse-height distribution of the MCP and, therefore, MCP gain. This proposal executes the same steps as the STIS MAMA Fold Distribution, Proposal 13149, as Cycle 20.

  4. STIS MAMA Fold Distribution

    NASA Astrophysics Data System (ADS)

    Wheeler, Thomas

    2011-10-01

    The performance of MAMA microchannel plates can be monitored using a MAMA fold analysis procedure. The fold analysis provides a measurement of the distribution of charge cloud sizes incident upon the anode giving some measure of changes in the pulse-height distribution of the MCP and, therefore, MCP gain. This proposal executes the same steps as the STIS MAMA Fold Analysis, Proposal 12416, as Cycle 18.

  5. A galaxy of folds.

    PubMed

    Alva, Vikram; Remmert, Michael; Biegert, Andreas; Lupas, Andrei N; Söding, Johannes

    2010-01-01

    Many protein classification systems capture homologous relationships by grouping domains into families and superfamilies on the basis of sequence similarity. Superfamilies with similar 3D structures are further grouped into folds. In the absence of discernable sequence similarity, these structural similarities were long thought to have originated independently, by convergent evolution. However, the growth of databases and advances in sequence comparison methods have led to the discovery of many distant evolutionary relationships that transcend the boundaries of superfamilies and folds. To investigate the contributions of convergent versus divergent evolution in the origin of protein folds, we clustered representative domains of known structure by their sequence similarity, treating them as point masses in a virtual 2D space which attract or repel each other depending on their pairwise sequence similarities. As expected, families in the same superfamily form tight clusters. But often, superfamilies of the same fold are linked with each other, suggesting that the entire fold evolved from an ancient prototype. Strikingly, some links connect superfamilies with different folds. They arise from modular peptide fragments of between 20 and 40 residues that co-occur in the connected folds in disparate structural contexts. These may be descendants of an ancestral pool of peptide modules that evolved as cofactors in the RNA world and from which the first folded proteins arose by amplification and recombination. Our galaxy of folds summarizes, in a single image, most known and many yet undescribed homologous relationships between protein superfamilies, providing new insights into the evolution of protein domains. PMID:19937658

  6. Effects of Knots on Protein Folding Properties

    PubMed Central

    Soler, Miguel A.; Faísca, Patrícia F. N.

    2013-01-01

    This work explores the impact of knots, knot depth and motif of the threading terminus in protein folding properties (kinetics, thermodynamics and mechanism) via extensive Monte Carlo simulations of lattice models. A knotted backbone has no effect on protein thermodynamic stability but it may affect key aspects of folding kinetics. In this regard, we found clear evidence for a functional advantage of knots: knots enhance kinetic stability because a knotted protein unfolds at a distinctively slower rate than its unknotted counterpart. However, an increase in knot deepness does not necessarily lead to more effective changes in folding properties. In this regard, a terminus with a non-trivial conformation (e.g. hairpin) can have a more dramatic effect in enhancing kinetic stability than knot depth. Nevertheless, our results suggest that the probability of the denatured ensemble to keep knotted is higher for proteins with deeper knots, indicating that knot depth plays a role in determining the topology of the denatured state. Refolding simulations starting from denatured knotted conformations show that not every knot is able to nucleate folding and further indicate that the formation of the knotting loop is a key event in the folding of knotted trefoils. They also show that there are specific native contacts within the knotted core that are crucial to keep a native knotting loop in denatured conformations which otherwise have no detectable structure. The study of the knotting mechanism reveals that the threading of the knotting loop generally occurs towards late folding in conformations that exhibit a significant degree of structural consolidation. PMID:24023962

  7. Unfolding Kinetics of the Human Telomere i-Motif Under a 10 pN Force Imposed by the α-Hemolysin Nanopore Identify Transient Folded-State Lifetimes at Physiological pH.

    PubMed

    Ding, Yun; Fleming, Aaron M; He, Lidong; Burrows, Cynthia J

    2015-07-22

    Cytosine (C)-rich DNA can adopt i-motif folds under acidic conditions, with the human telomere i-motif providing a well-studied example. The dimensions of this i-motif are appropriate for capture in the nanocavity of the α-hemolysin (α-HL) protein pore under an electrophoretic force. Interrogation of the current vs time (i-t) traces when the i-motif interacts with α-HL identified characteristic signals that were pH dependent. These features were evaluated from pH 5.0 to 7.2, a region surrounding the transition pH of the i-motif (6.1). When the i-motif without polynucleotide tails was studied at pH 5.0, the folded structure entered the nanocavity of α-HL from either the top or bottom face to yield characteristic current patterns. Addition of a 5' 25-mer poly-2'-deoxyadensosine tail allowed capture of the i-motif from the unfolded terminus, and this was used to analyze the pH dependency of unfolding. At pH values below the transition point, only folded strands were observed, and when the pH was increased above the transition pH, the number of folded events decreased, while the unfolded events increased. At pH 6.8 and 7.2 4% and 2% of the strands were still folded, respectively. The lifetimes for the folded states at pH 6.8 and 7.2 were 21 and 9 ms, respectively, at 160 mV electrophoretic force. These lifetimes are sufficiently long to affect enzymes operating on DNA. Furthermore, these transient lifetimes are readily obtained using the α-HL nanopore, a feature that is not easily achievable by other methods.

  8. Mathematics Through Paper Folding.

    ERIC Educational Resources Information Center

    Olson, Alton T.

    This booklet is a revised edition of Donovan Johnson's "Paper Folding for the Mathematics Class" (ED 077 711). It begins with directions for folding basic constructions such as as a straight line, the line perpendicular to a given line passing through a given point, and the bisector of an angle. Subsequent chapters cover concepts related to…

  9. Folding by Design

    NASA Astrophysics Data System (ADS)

    Dodd, Paul; Damasceno, Pablo; Glotzer, Sharon

    2014-03-01

    A form of self-assembly, ``self-folding'' presents an alternative approach to the creation of reconfigurable, responsive materials with applications ranging from robotics to drug design. However, the complexity of interactions present in biological and engineered systems that undergo folding makes it challenging to isolate the main factors controlling their assembly and dis-assembly. Here we use computer simulations of simple, minimalistic self-foldable structures and investigate their stochastic folding process. By dynamically accessing all the states that lead to, or inhibit, successful folding, we show that the mechanisms by which general stochastic systems can achieve their ``native'' structures can be identified and used to design rules for optimized folding propensity. Research supported by the National Science Foundation, Emerging Frontiers in Research and Innovation Award # EFRI-1240264.

  10. Circuit topology of self-interacting chains: implications for folding and unfolding dynamics.

    PubMed

    Mugler, Andrew; Tans, Sander J; Mashaghi, Alireza

    2014-11-01

    Understanding the relationship between molecular structure and folding is a central problem in disciplines ranging from biology to polymer physics and DNA origami. Topology can be a powerful tool to address this question. For a folded linear chain, the arrangement of intra-chain contacts is a topological property because rearranging the contacts requires discontinuous deformations. Conversely, the topology is preserved when continuously stretching the chain while maintaining the contact arrangement. Here we investigate how the folding and unfolding of linear chains with binary contacts is guided by the topology of contact arrangements. We formalize the topology by describing the relations between any two contacts in the structure, which for a linear chain can either be in parallel, in series, or crossing each other. We show that even when other determinants of folding rate such as contact order and size are kept constant, this 'circuit' topology determines folding kinetics. In particular, we find that the folding rate increases with the fractions of parallel and crossed relations. Moreover, we show how circuit topology constrains the conformational phase space explored during folding and unfolding: the number of forbidden unfolding transitions is found to increase with the fraction of parallel relations and to decrease with the fraction of series relations. Finally, we find that circuit topology influences whether distinct intermediate states are present, with crossed contacts being the key factor. The approach presented here can be more generally applied to questions on molecular dynamics, evolutionary biology, molecular engineering, and single-molecule biophysics.

  11. Quantum theory on protein folding

    NASA Astrophysics Data System (ADS)

    Luo, LiaoFu

    2014-03-01

    The conformational change of biological macromolecule is investigated from the point of quantum transition. A quantum theory on protein folding is proposed. Compared with other dynamical variables such as mobile electrons, chemical bonds and stretching-bending vibrations the molecular torsion has the lowest energy and can be looked as the slow variable of the system. Simultaneously, from the multi-minima property of torsion potential the local conformational states are well defined. Following the idea that the slow variables slave the fast ones and using the nonadiabaticity operator method we deduce the Hamiltonian describing conformational change. It is shown that the influence of fast variables on the macromolecule can fully be taken into account through a phase transformation of slow variable wave function. Starting from the conformation-transition Hamiltonian the nonradiative matrix element was calculated and a general formulas for protein folding rate was deduced. The analytical form of the formula was utilized to study the temperature dependence of protein folding rate and the curious non-Arrhenius temperature relation was interpreted. By using temperature dependence data the multi-torsion correlation was studied. The decoherence time of quantum torsion state is estimated. The proposed folding rate formula gives a unifying approach for the study of a large class problems of biological conformational change.

  12. Chemical and Biological Kinetics

    NASA Astrophysics Data System (ADS)

    Emanuel', N. M.

    1981-10-01

    Examples of the application of the methods and ideas of chemical kinetics in various branches of chemistry and biology are considered and the results of studies on the kinetics and mechanisms of autoxidation and inhibited and catalysed oxidation of organic substances in the liquid phase are surveyed. Problems of the kinetics of the ageing of polymers and the principles of their stabilisation are discussed and certain trends in biological kinetics (kinetics of tumour growth, kinetic criteria of the effectiveness of chemotherapy, problems of gerontology, etc.) are considered. The bibliography includes 281 references.

  13. On the Role of Entropy in the Protein Folding Process

    NASA Astrophysics Data System (ADS)

    Hoppe, Travis

    2011-12-01

    A protein's ultimate function and activity is determined by the unique three-dimensional structure taken by the folding process. Protein malfunction due to misfolding is the culprit of many clinical disorders, such as abnormal protein aggregations. This leads to neurodegenerative disorders like Huntington's and Alzheimer's disease. We focus on a subset of the folding problem, exploring the role and effects of entropy on the process of protein folding. Four major concepts and models are developed and each pertains to a specific aspect of the folding process: entropic forces, conformational states under crowding, aggregation, and macrostate kinetics from microstate trajectories. The exclusive focus on entropy is well-suited for crowding studies, as many interactions are nonspecific. We show how a stabilizing entropic force can arise purely from the motion of crowders in solution. In addition we are able to make a a quantitative prediction of the crowding effect with an implicit crowding approximation using an aspherical scaled-particle theory. In order to investigate the effects of aggregation, we derive a new operator expansion method to solve the Ising/Potts model with external fields over an arbitrary graph. Here the external fields are representative of the entropic forces. We show that this method reduces the problem of calculating the partition function to the solution of recursion relations. Many of the methods employed are coarse-grained approximations. As such, it is useful to have a viable method for extracting macrostate information from time series data. We develop a method to cluster the microstates into physically meaningful macrostates by grouping similar relaxation times from a transition matrix. Overall, the studied topics allow us to understand deeper the complicated process involving proteins.

  14. NUV MAMA Fold Distribution

    NASA Astrophysics Data System (ADS)

    Wheeler, Thomas

    2009-07-01

    The performance of MAMA microchannel plate can be monitored using a MAMA fold analysis procedure. The fold analysis provides a measurement of the distribution of charge cloud sizes incident upon the anode giving some measure of changes in the pulse-height distribution of the MCP and, therefore, MCP gain. This proposal executes the same steps as the COS SMOV as proposal 13555 {visit 5}.

  15. Learning Protein Folding Energy Functions

    PubMed Central

    Guan, Wei; Ozakin, Arkadas; Gray, Alexander; Borreguero, Jose; Pandit, Shashi; Jagielska, Anna; Wroblewska, Liliana; Skolnick, Jeffrey

    2014-01-01

    A critical open problem in ab initio protein folding is protein energy function design, which pertains to defining the energy of protein conformations in a way that makes folding most efficient and reliable. In this paper, we address this issue as a weight optimization problem and utilize a machine learning approach, learning-to-rank, to solve this problem. We investigate the ranking-via-classification approach, especially the RankingSVM method and compare it with the state-of-the-art approach to the problem using the MINUIT optimization package. To maintain the physicality of the results, we impose non-negativity constraints on the weights. For this we develop two efficient non-negative support vector machine (NNSVM) methods, derived from L2-norm SVM and L1-norm SVMs, respectively. We demonstrate an energy function which maintains the correct ordering with respect to structure dissimilarity to the native state more often, is more efficient and reliable for learning on large protein sets, and is qualitatively superior to the current state-of-the-art energy function. PMID:25311546

  16. Folding without charges

    PubMed Central

    Kurnik, Martin; Hedberg, Linda; Danielsson, Jens; Oliveberg, Mikael

    2012-01-01

    Surface charges of proteins have in several cases been found to function as “structural gatekeepers,” which avoid unwanted interactions by negative design, for example, in the control of protein aggregation and binding. The question is then if side-chain charges, due to their desolvation penalties, play a corresponding role in protein folding by avoiding competing, misfolded traps? To find out, we removed all 32 side-chain charges from the 101-residue protein S6 from Thermus thermophilus. The results show that the charge-depleted S6 variant not only retains its native structure and cooperative folding transition, but folds also faster than the wild-type protein. In addition, charge removal unleashes pronounced aggregation on longer timescales. S6 provides thus an example where the bias toward native contacts of a naturally evolved protein sequence is independent of charges, and point at a fundamental difference in the codes for folding and intermolecular interaction: specificity in folding is governed primarily by hydrophobic packing and hydrogen bonding, whereas solubility and binding relies critically on the interplay of side-chain charges. PMID:22454493

  17. Accurate prediction of cellular co-translational folding indicates proteins can switch from post- to co-translational folding

    NASA Astrophysics Data System (ADS)

    Nissley, Daniel A.; Sharma, Ajeet K.; Ahmed, Nabeel; Friedrich, Ulrike A.; Kramer, Günter; Bukau, Bernd; O'Brien, Edward P.

    2016-02-01

    The rates at which domains fold and codons are translated are important factors in determining whether a nascent protein will co-translationally fold and function or misfold and malfunction. Here we develop a chemical kinetic model that calculates a protein domain's co-translational folding curve during synthesis using only the domain's bulk folding and unfolding rates and codon translation rates. We show that this model accurately predicts the course of co-translational folding measured in vivo for four different protein molecules. We then make predictions for a number of different proteins in yeast and find that synonymous codon substitutions, which change translation-elongation rates, can switch some protein domains from folding post-translationally to folding co-translationally--a result consistent with previous experimental studies. Our approach explains essential features of co-translational folding curves and predicts how varying the translation rate at different codon positions along a transcript's coding sequence affects this self-assembly process.

  18. Folding pathways of the Tetrahymena ribozyme.

    PubMed

    Mitchell, David; Russell, Rick

    2014-06-12

    Like many structured RNAs, the Tetrahymena group I intron ribozyme folds through multiple pathways and intermediates. Under standard conditions in vitro, a small fraction reaches the native state (N) with kobs ≈ 0.6 min(-1), while the remainder forms a long-lived misfolded conformation (M) thought to differ in topology. These alternative outcomes reflect a pathway that branches late in folding, after disruption of a trapped intermediate (Itrap). Here we use catalytic activity to probe the folding transitions from Itrap to the native and misfolded states. We show that mutations predicted to weaken the core helix P3 do not increase the rate of folding from Itrap but they increase the fraction that reaches the native state rather than forming the misfolded state. Thus, P3 is disrupted during folding to the native state but not to the misfolded state, and P3 disruption occurs after the rate-limiting step. Interestingly, P3-strengthening mutants also increase native folding. Additional experiments show that these mutants are rapidly committed to folding to the native state, although they reach the native state with approximately the same rate constant as the wild-type ribozyme (~1 min(-1)). Thus, the P3-strengthening mutants populate a distinct pathway that includes at least one intermediate but avoids the M state, most likely because P3 and the correct topology are formed early. Our results highlight multiple pathways in RNA folding and illustrate how kinetic competitions between rapid events can have long-lasting effects because the "choice" is enforced by energy barriers that grow larger as folding progresses.

  19. Folding pathways of the Tetrahymena ribozyme

    PubMed Central

    Mitchell, David; Russell, Rick

    2014-01-01

    Like many structured RNAs, the Tetrahymena group I intron ribozyme folds through multiple pathways and intermediates. Under standard conditions in vitro, a small fraction reaches the native state (N) with kobs ≈ 0.6 min–1, while the remainder forms a long-lived misfolded conformation (M) thought to differ in topology. These alternative outcomes reflect a pathway that branches late in folding, after disruption of a trapped intermediate (Itrap). Here, we use catalytic activity to probe the folding transitions from Itrap to the native and misfolded states. We show that mutations predicted to weaken the core helix P3 do not increase the rate of folding from Itrap but they increase the fraction that reaches the native state rather than forming the misfolded state. Thus, P3 is disrupted during folding to the native state but not to the misfolded state, and P3 disruption occurs after the rate-limiting step. Interestingly, P3-strengthening mutants also increase native folding. Additional experiments show that these mutants are rapidly committed to folding to the native state, although they reach the native state with approximately the same rate constant as the wild-type ribozyme (~1 min–1). Thus, the P3-strengthening mutants populate a distinct pathway that includes at least one intermediate but avoids the M state, most likely because P3 and the correct topology are formed early. Our results highlight multiple pathways in RNA folding and illustrate how kinetic competitions between rapid events can have long-lasting effects because the ‘choice’ is enforced by energy barriers that grow larger as folding progresses. PMID:24747051

  20. Folding a protein by discretizing its backbone torsional dynamics

    NASA Astrophysics Data System (ADS)

    Fernández, Ariel

    1999-05-01

    The aim of this work is to provide a coarse codification of local conformational constraints associated with each folding motif of a peptide chain in order to obtain a rough solution to the protein folding problem. This is accomplished by implementing a discretized version of the soft-mode dynamics on a personal computer (PC). Our algorithm mimics a parallel process as it evaluates concurrent folding possibilities by pattern recognition. It may be implemented in a PC as a sequence of perturbation-translation-renormalization (p-t-r) cycles performed on a matrix of local topological constraints (LTM). This requires suitable representational tools and a periodic quenching of the dynamics required for renormalization. We introduce a description of the peptide chain based on a local discrete variable the values of which label the basins of attraction of the Ramachandran map for each residue. Thus, the local variable indicates the basin in which the torsional coordinates of each residue lie at a given time. In addition, a coding of local topological constraints associated with each secondary and tertiary structural motif is introduced. Our treatment enables us to adopt a computation time step of 81 ps, a value far larger than hydrodynamic drag time scales. Folding pathways are resolved as transitions between patterns of locally encoded structural signals that change within the 10 μs-100 ms time scale range. These coarse folding pathways are generated by the periodic search for structural patterns in the time-evolving LTM. Each pattern is recorded as a contact matrix, an operation subject to a renormalization feedback loop. The validity of our approach is tested vis-a-vis experimentally-probed folding pathways eventually generating tertiary interactions in proteins which recover their active structure under in vitro renaturation conditions. As an illustration, we focus on determining significant folding intermediates and late kinetic bottlenecks that occur within the

  1. Probing the cytochrome c' folding landscape.

    PubMed

    Pletneva, Ekaterina V; Zhao, Ziqing; Kimura, Tetsunari; Petrova, Krastina V; Gray, Harry B; Winkler, Jay R

    2007-11-01

    The folding kinetics of R. palustris cytochrome c' (cyt c') have been monitored by heme absorption and native Trp72 fluorescence at pH 5. The Trp72 fluorescence burst signal suggests early compaction of the polypeptide ensemble. Analysis of heme transient absorption spectra reveals deviations from two-state behavior, including a prominent slow phase that is accelerated by the prolyl isomerase cyclophilin. A nonnative proline configuration (Pro21) likely interferes with the formation of the helical bundle surrounding the heme.

  2. Steric constraints as folding coadjuvant

    NASA Astrophysics Data System (ADS)

    Tarragó, M. E.; Rocha, Luiz F.; Dasilva, R. A.; Caliri, A.

    2003-03-01

    Through the analyses of the Miyazawa-Jernigan matrix it has been shown that the hydrophobic effect generates the dominant driving force for protein folding. By using both lattice and off-lattice models, it is shown that hydrophobic-type potentials are indeed efficient in inducing the chain through nativelike configurations, but they fail to provide sufficient stability so as to keep the chain in the native state. However, through comparative Monte Carlo simulations, it is shown that hydrophobic potentials and steric constraints are two basic ingredients for the folding process. Specifically, it is shown that suitable pairwise steric constraints introduce strong changes on the configurational activity, whose main consequence is a huge increase in the overall stability condition of the native state; detailed analysis of the effects of steric constraints on the heat capacity and configurational activity are provided. The present results support the view that the folding problem of globular proteins can be approached as a process in which the mechanism to reach the native conformation and the requirements for the globule stability are uncoupled.

  3. Visualizing chaperone-assisted protein folding.

    PubMed

    Horowitz, Scott; Salmon, Loïc; Koldewey, Philipp; Ahlstrom, Logan S; Martin, Raoul; Quan, Shu; Afonine, Pavel V; van den Bedem, Henry; Wang, Lili; Xu, Qingping; Trievel, Raymond C; Brooks, Charles L; Bardwell, James C A

    2016-07-01

    Challenges in determining the structures of heterogeneous and dynamic protein complexes have greatly hampered past efforts to obtain a mechanistic understanding of many important biological processes. One such process is chaperone-assisted protein folding. Obtaining structural ensembles of chaperone-substrate complexes would ultimately reveal how chaperones help proteins fold into their native state. To address this problem, we devised a new structural biology approach based on X-ray crystallography, termed residual electron and anomalous density (READ). READ enabled us to visualize even sparsely populated conformations of the substrate protein immunity protein 7 (Im7) in complex with the Escherichia coli chaperone Spy, and to capture a series of snapshots depicting the various folding states of Im7 bound to Spy. The ensemble shows that Spy-associated Im7 samples conformations ranging from unfolded to partially folded to native-like states and reveals how a substrate can explore its folding landscape while being bound to a chaperone. PMID:27239796

  4. Folds and Etudes

    ERIC Educational Resources Information Center

    Bean, Robert

    2007-01-01

    In this article, the author talks about "Folds" and "Etudes" which are images derived from anonymous typing exercises that he found in a used copy of "Touch Typing Made Simple". "Etudes" refers to the musical tradition of studies for a solo instrument, which is a typewriter. Typing exercises are repetitive attempts to type words and phrases…

  5. Simultaneous Alignment and Folding of Protein Sequences

    PubMed Central

    Waldispühl, Jérôme; O'Donnell, Charles W.; Will, Sebastian; Devadas, Srinivas; Backofen, Rolf

    2014-01-01

    Abstract Accurate comparative analysis tools for low-homology proteins remains a difficult challenge in computational biology, especially sequence alignment and consensus folding problems. We present partiFold-Align, the first algorithm for simultaneous alignment and consensus folding of unaligned protein sequences; the algorithm's complexity is polynomial in time and space. Algorithmically, partiFold-Align exploits sparsity in the set of super-secondary structure pairings and alignment candidates to achieve an effectively cubic running time for simultaneous pairwise alignment and folding. We demonstrate the efficacy of these techniques on transmembrane β-barrel proteins, an important yet difficult class of proteins with few known three-dimensional structures. Testing against structurally derived sequence alignments, partiFold-Align significantly outperforms state-of-the-art pairwise and multiple sequence alignment tools in the most difficult low-sequence homology case. It also improves secondary structure prediction where current approaches fail. Importantly, partiFold-Align requires no prior training. These general techniques are widely applicable to many more protein families (partiFold-Align is available at http://partifold.csail.mit.edu/). PMID:24766258

  6. Design principles for rapid folding of knotted DNA nanostructures.

    PubMed

    Kočar, Vid; Schreck, John S; Čeru, Slavko; Gradišar, Helena; Bašić, Nino; Pisanski, Tomaž; Doye, Jonathan P K; Jerala, Roman

    2016-01-01

    Knots are some of the most remarkable topological features in nature. Self-assembly of knotted polymers without breaking or forming covalent bonds is challenging, as the chain needs to be threaded through previously formed loops in an exactly defined order. Here we describe principles to guide the folding of highly knotted single-chain DNA nanostructures as demonstrated on a nano-sized square pyramid. Folding of knots is encoded by the arrangement of modules of different stability based on derived topological and kinetic rules. Among DNA designs composed of the same modules and encoding the same topology, only the one with the folding pathway designed according to the 'free-end' rule folds efficiently into the target structure. Besides high folding yield on slow annealing, this design also folds rapidly on temperature quenching and dilution from chemical denaturant. This strategy could be used to design folding of other knotted programmable polymers such as RNA or proteins.

  7. Design principles for rapid folding of knotted DNA nanostructures

    PubMed Central

    Kočar, Vid; Schreck, John S.; Čeru, Slavko; Gradišar, Helena; Bašić, Nino; Pisanski, Tomaž; Doye, Jonathan P. K.; Jerala, Roman

    2016-01-01

    Knots are some of the most remarkable topological features in nature. Self-assembly of knotted polymers without breaking or forming covalent bonds is challenging, as the chain needs to be threaded through previously formed loops in an exactly defined order. Here we describe principles to guide the folding of highly knotted single-chain DNA nanostructures as demonstrated on a nano-sized square pyramid. Folding of knots is encoded by the arrangement of modules of different stability based on derived topological and kinetic rules. Among DNA designs composed of the same modules and encoding the same topology, only the one with the folding pathway designed according to the ‘free-end' rule folds efficiently into the target structure. Besides high folding yield on slow annealing, this design also folds rapidly on temperature quenching and dilution from chemical denaturant. This strategy could be used to design folding of other knotted programmable polymers such as RNA or proteins. PMID:26887681

  8. Modelling of lateral fold growth and fold linkage: Applications to fold-and-thrust belt tectonics

    NASA Astrophysics Data System (ADS)

    Grasemann, Bernhard; Schmalholz, Stefan

    2013-04-01

    We use a finite element model to investigate the three-dimensional fold growth and interference of two initially isolated fold segments. The most critical parameter, which controls the fold linkage mode, is the phase difference between the laterally growing fold hinge lines: 1) "Linear-linkage" yields a sub-cylindrical fold with a saddle at the location where the two initial folds linked. 2) "Oblique-linkage" produces a curved fold resembling a Type II refold structure. 3) "Oblique-no-linkage" results in two curved folds with fold axes plunging in opposite directions. 4) "Linear-no-linkage" yields a fold train of two separate sub-cylindrical folds with fold axes plunging in opposite directions. The transition from linkage to no-linkage occurs when the fold separation between the initially isolated folds is slightly larger than one half of the low-amplitude fold wavelength. The model results compare well with previously published plasticine analogue models and can be directly applied to the investigation of fold growth history in fold-and-thust belts. An excellent natural example of lateral fold linkage is described from the Zagros fold-and-thrust belt in the Kurdistan Region of Iraq. The fold growth in this region is not controlled by major thrust faults but the shortening of the Paleozoic to Cenozoic passive margin sediments of the Arabian plate occurred mainly by detachment folding. The sub-cylindrical anticlines with hinge-parallel lengths of more than 50 km have not developed from single sub-cylindrical embryonic folds but they have merged from different fold segments that joined laterally during fold amplification and lateral fold growth. Linkage points are marked by geomorphological saddle points which are structurally the lowermost points of antiforms and points of principal curvatures with opposite sign. Linkage points can significantly influence the migration of mineral-rich fluids and hydrocarbons and are therefore of great economic importance.

  9. Oxidative folding: recent developments.

    PubMed

    Szarka, András; Bánhegyi, Gábor

    2011-10-01

    Disulfide bond formation in proteins is an effective tool of both structure stabilization and redox regulation. The prokaryotic periplasm and the endoplasmic reticulum of eukaryotes were long considered as the only compartments for enzyme mediated formation of stable disulfide bonds. Recently, the mitochondrial intermembrane space has emerged as the third protein-oxidizing compartment. The classic view on the mechanism of oxidative folding in the endoplasmic reticulum has also been reshaped by new observations. Moreover, besides the structure stabilizing function, reversible disulfide bridge formation in some proteins of the endoplasmic reticulum, seems to play a regulatory role. This review briefly summarizes the present knowledge of the redox systems supporting oxidative folding, emphasizing recent developments. PMID:25962043

  10. The protein folding network

    NASA Astrophysics Data System (ADS)

    Rao, Francesco; Caflisch, Amedeo

    2004-03-01

    Networks are everywhere. The conformation space of a 20-residue antiparallel beta-sheet peptide [1], sampled by molecular dynamics simulations, is mapped to a network. Conformations are nodes of the network, and the transitions between them are links. As previously found for the World-Wide Web as well as for social and biological networks , the conformation space contains highly connected hubs like the native state which is the most populated free energy basin. Furthermore, the network shows a hierarchical modularity [2] which is consistent with the funnel mechanism of folding [3] and is not observed for a random heteropolymer lacking a native state. Here we show that the conformation space network describes the free energy landscape without requiring projections into arbitrarily chosen reaction coordinates. The network analysis provides a basis for understanding the heterogeneity of the folding transition state and the existence of multiple pathways. [1] P. Ferrara and A. Caflisch, Folding simulations of a three-stranded antiparallel beta-sheet peptide, PNAS 97, 10780-10785 (2000). [2] Ravasz, E. and Barabási, A. L. Hierarchical organization in complex networks. Phys. Rev. E 67, 026112 (2003). [3] Dill, K. and Chan, H From Levinthal to pathways to funnels. Nature Struct. Biol. 4, 10-19 (1997)

  11. Folded waveguide coupler

    DOEpatents

    Owens, Thomas L.

    1988-03-01

    A resonant cavity waveguide coupler for ICRH of a magnetically confined plasma. The coupler consists of a series of inter-leaved metallic vanes disposed withn an enclosure analogous to a very wide, simple rectangular waveguide that has been "folded" several times. At the mouth of the coupler, a polarizing plate is provided which has coupling apertures aligned with selected folds of the waveguide through which rf waves are launched with magnetic fields of the waves aligned in parallel with the magnetic fields confining the plasma being heated to provide coupling to the fast magnetosonic wave within the plasma in the frequency usage of from about 50-200 mHz. A shorting plate terminates the back of the cavity at a distance approximately equal to one-half the guide wavelength from the mouth of the coupler to ensure that the electric field of the waves launched through the polarizing plate apertures are small while the magnetic field is near a maximum. Power is fed into the coupler folded cavity by means of an input coaxial line feed arrangement at a point which provides an impedance match between the cavity and the coaxial input line.

  12. Ab initio RNA folding

    NASA Astrophysics Data System (ADS)

    Cragnolini, Tristan; Derreumaux, Philippe; Pasquali, Samuela

    2015-06-01

    RNA molecules are essential cellular machines performing a wide variety of functions for which a specific three-dimensional structure is required. Over the last several years, the experimental determination of RNA structures through x-ray crystallography and NMR seems to have reached a plateau in the number of structures resolved each year, but as more and more RNA sequences are being discovered, the need for structure prediction tools to complement experimental data is strong. Theoretical approaches to RNA folding have been developed since the late nineties, when the first algorithms for secondary structure prediction appeared. Over the last 10 years a number of prediction methods for 3D structures have been developed, first based on bioinformatics and data-mining, and more recently based on a coarse-grained physical representation of the systems. In this review we are going to present the challenges of RNA structure prediction and the main ideas behind bioinformatic approaches and physics-based approaches. We will focus on the description of the more recent physics-based phenomenological models and on how they are built to include the specificity of the interactions of RNA bases, whose role is critical in folding. Through examples from different models, we will point out the strengths of physics-based approaches, which are able not only to predict equilibrium structures, but also to investigate dynamical and thermodynamical behavior, and the open challenges to include more key interactions ruling RNA folding.

  13. Macrotransport-solidification kinetics modeling of equiaxed dendritic growth: Part II. Computation problems and validation on INCONEL 718 superalloy castings

    NASA Astrophysics Data System (ADS)

    Nastac, L.; Stefanescu, D. M.

    1996-12-01

    In Part I of the article, a new analytical model that describes solidification of equiaxed dendrites was presented. In this part of the article, the model is used to simulate the solidification of INCONEL 718 superalloy castings. The model was incorporated into a commercial finite-element code, PROCAST. A special procedure called microlatent heat method (MLHM) was used for coupling between macroscopic heat flow and microscopic growth kinetics. A criterion for time-stepping selection in microscopic modeling has been derived in conjunction with MLHM. Reductions in computational (CPU) time up to 90 pct over the classic latent heat method were found by adopting this coupling. Validation of the model was performed against experimental data for an INCONEL 718 superalloy casting. In the present calculations, the model for globulitic dendrite was used. The evolution of fraction of solid calculated with the present model was compared with Scheil’s model and experiments. An important feature in solidification of INCONEL 718 is the detrimental Laves phase. Laves phase content is directly related to the intensity of microsegregation of niobium, which is very sensitive to the evolution of the fraction of solid. It was found that there is a critical cooling rate at which the amount of Laves phase is maximum. The critical cooling rate is not a function of material parameters (diffusivity, partition coefficient, etc.). It depends only on the grain size and solidification time. The predictions generated with the present model are shown to agree very well with experiments.

  14. Folding funnels, binding funnels, and protein function.

    PubMed Central

    Tsai, C. J.; Kumar, S.; Ma, B.; Nussinov, R.

    1999-01-01

    , with a range of complexed conformations. Hence, knowledge of the shape of the folding funnels is biologically very useful. The converse also holds: If kinetic and thermodynamic data are available, hints regarding the role of the protein and its binding selectivity may be obtained. Thus, the utility of the concept of the funnel carries over to the origin of the protein and to its function. PMID:10386868

  15. Folding of Pollen Grains

    NASA Astrophysics Data System (ADS)

    Katifori, Eleni; Alben, Silas; Cerda, Enrique; Nelson, David; Dumais, Jacques

    2008-03-01

    At dehiscence, which occurs when the anther reaches maturity and opens, pollen grains dehydrate and their volume is reduced. The pollen wall deforms to accommodate the volume loss, and the deformation pathway depends on the initial turgid pollen grain geometry and the mechanical properties of the pollen wall. We demonstrate, using both experimental and theoretical approaches, that the design of the apertures (areas on the pollen wall where the stretching and the bending modulus are reduced) is critical for controlling the folding pattern, and ensures the pollen grain viability. An excellent fit to the experiments is obtained using a discretized version of the theory of thin elastic shells.

  16. Cellular Mechanisms of Membrane Protein Folding

    PubMed Central

    Skach, William R.

    2010-01-01

    The membrane protein folding problem can be articulated by two central questions. How is protein topology established by selective peptide transport to opposite sides of the cellular membrane? And how are transmembrane segments inserted, integrated and folded within the lipid bilayer? In eukaryotes, this process usually takes place in the endoplasmic reticulum (ER) coincident with protein synthesis, and is facilitated by the translating Ribosome and the Sec61 Translocon Complex (RTC). At its core, the RTC forms a dynamic pathway through which the elongating nascent polypeptide moves as it is delivered into cytosolic, lumenal and lipid compartments. This perspective will focus on emerging evidence that the RTC functions as a protein folding machine that restricts conformational space by establishing transmembrane topology and yet provides a permissive environment that enables nascent transmembrane domains to efficiently progress down their folding energy landscape. PMID:19491932

  17. Fragility of Liquids, Polyamorphism, Nucleation, and Folding Directions, in the Landscape Paradigm

    NASA Astrophysics Data System (ADS)

    Angell, C. A.

    1998-03-01

    folding problem. The possibility exists that in certain cases an aberrant step in the nucleation event, facilitated by mutant nucleotide sequences or by third agents (heterogeneous nucleating agents), will trigger folding down an alternative and pathogenic route to a second stable state. This possibility should be evaluated, using nucleation kinetics analysis techniques, as an approach to understanding the initiation of ``mad cow" disease cerebral pathology.

  18. Protein photo-folding and quantum folding theory.

    PubMed

    Luo, Liaofu

    2012-06-01

    The rates of protein folding with photon absorption or emission and the cross section of photon -protein inelastic scattering are calculated from quantum folding theory by use of a field-theoretical method. All protein photo-folding processes are compared with common protein folding without the interaction of photons (non-radiative folding). It is demonstrated that there exists a common factor (thermo-averaged overlap integral of the vibration wave function, TAOI) for protein folding and protein photo-folding. Based on this finding it is predicted that (i) the stimulated photo-folding rates and the photon-protein resonance Raman scattering sections show the same temperature dependence as protein folding; (ii) the spectral line of the electronic transition is broadened to a band that includes an abundant vibration spectrum without and with conformational transitions, and the width of each vibration spectral line is largely reduced. The particular form of the folding rate-temperature relation and the abundant spectral structure imply the existence of quantum tunneling between protein conformations in folding and photo-folding that demonstrates the quantum nature of the motion of the conformational-electronic system.

  19. Folded pendulum tiltmeter

    NASA Astrophysics Data System (ADS)

    Wu, Shuchao; Fan, Shuhua; Luo, Jun; Hsu, Houtse

    2002-05-01

    The application of the folded pendulum (FP) as a tiltmeter is proposed and some features of it have been studied both theoretically and experimentally. First, FP could have a quite low resonance frequency due to its mechanical structure. The period of our prototype FP is 6.2 s and the amplification factor is about 100. Second, FP is not sensitive to the environmental temperature variation but seriously affected by the temperature gradient. The experiment with a temperature gradient modulation shows that the static equilibrium position of the FP will change by 2.5 μrad if the temperature difference between the horizontal platform and the base of the FP is 0.1 °C. With a prototype FP, we have observed obvious tilt tides, and the calibration result shows that the resolution of the FP is about 1.2 nrad.

  20. How the diffusivity profile reduces the arbitrariness of protein folding free energies

    NASA Astrophysics Data System (ADS)

    Hinczewski, M.; von Hansen, Y.; Dzubiella, J.; Netz, R. R.

    2010-06-01

    The concept of a protein diffusing in its free-energy folding landscape has been fruitful for both theory and experiment. Yet the choice of the reaction coordinate (RC) introduces an undesirable degree of arbitrariness into the problem. We analyze extensive simulation data of an α-helix in explicit water solvent as it stochastically folds and unfolds. The free-energy profiles for different RCs exhibit significant variations, some having an activation barrier, while others not. We show that this variation has little effect on the predicted folding kinetics if the diffusivity profiles are properly taken into account. This kinetic quasi-universality is rationalized by an RC rescaling, which, due to the reparameterization invariance of the Fokker-Planck equation, allows the combination of free-energy and diffusivity effects into a single function, the rescaled free-energy profile. This rescaled free energy indeed shows less variation among different RCs than the bare free energy and diffusivity profiles separately do, if we properly distinguish between RCs that contain knowledge of the native state and those that are purely geometric in nature. Our method for extracting diffusivity profiles is easily applied to experimental single molecule time series data and might help to reconcile conflicts that arise when comparing results from different experimental probes for the same protein.

  1. Protein folding and protein metallocluster studies using synchrotron small angler X-ray scattering

    SciTech Connect

    Eliezer, D.

    1994-06-01

    Proteins, biological macromolecules composed of amino-acid building blocks, possess unique three dimensional shapes or conformations which are intimately related to their biological function. All of the information necessary to determine this conformation is stored in a protein`s amino acid sequence. The problem of understanding the process by which nature maps protein amino-acid sequences to three-dimensional conformations is known as the protein folding problem, and is one of the central unsolved problems in biophysics today. The possible applications of a solution are broad, ranging from the elucidation of thousands of protein structures to the rational modification and design of protein-based drugs. The scattering of X-rays by matter has long been useful as a tool for the characterization of physical properties of materials, including biological samples. The high photon flux available at synchrotron X-ray sources allows for the measurement of scattering cross-sections of dilute and/or disordered samples. Such measurements do not yield the detailed geometrical information available from crystalline samples, but do allow for lower resolution studies of dynamical processes not observable in the crystalline state. The main focus of the work described here has been the study of the protein folding process using time-resolved small-angle x-ray scattering measurements. The original intention was to observe the decrease in overall size which must accompany the folding of a protein from an extended conformation to its compact native state. Although this process proved too fast for the current time-resolution of the technique, upper bounds were set on the probable compaction times of several small proteins. In addition, an interesting and unexpected process was detected, in which the folding protein passes through an intermediate state which shows a tendency to associate. This state is proposed to be a kinetic molten globule folding intermediate.

  2. Improving Protein Fold Recognition by Deep Learning Networks

    PubMed Central

    Jo, Taeho; Hou, Jie; Eickholt, Jesse; Cheng, Jianlin

    2015-01-01

    For accurate recognition of protein folds, a deep learning network method (DN-Fold) was developed to predict if a given query-template protein pair belongs to the same structural fold. The input used stemmed from the protein sequence and structural features extracted from the protein pair. We evaluated the performance of DN-Fold along with 18 different methods on Lindahl’s benchmark dataset and on a large benchmark set extracted from SCOP 1.75 consisting of about one million protein pairs, at three different levels of fold recognition (i.e., protein family, superfamily, and fold) depending on the evolutionary distance between protein sequences. The correct recognition rate of ensembled DN-Fold for Top 1 predictions is 84.5%, 61.5%, and 33.6% and for Top 5 is 91.2%, 76.5%, and 60.7% at family, superfamily, and fold levels, respectively. We also evaluated the performance of single DN-Fold (DN-FoldS), which showed the comparable results at the level of family and superfamily, compared to ensemble DN-Fold. Finally, we extended the binary classification problem of fold recognition to real-value regression task, which also show a promising performance. DN-Fold is freely available through a web server at http://iris.rnet.missouri.edu/dnfold. PMID:26634993

  3. Improving Protein Fold Recognition by Deep Learning Networks.

    PubMed

    Jo, Taeho; Hou, Jie; Eickholt, Jesse; Cheng, Jianlin

    2015-12-04

    For accurate recognition of protein folds, a deep learning network method (DN-Fold) was developed to predict if a given query-template protein pair belongs to the same structural fold. The input used stemmed from the protein sequence and structural features extracted from the protein pair. We evaluated the performance of DN-Fold along with 18 different methods on Lindahl's benchmark dataset and on a large benchmark set extracted from SCOP 1.75 consisting of about one million protein pairs, at three different levels of fold recognition (i.e., protein family, superfamily, and fold) depending on the evolutionary distance between protein sequences. The correct recognition rate of ensembled DN-Fold for Top 1 predictions is 84.5%, 61.5%, and 33.6% and for Top 5 is 91.2%, 76.5%, and 60.7% at family, superfamily, and fold levels, respectively. We also evaluated the performance of single DN-Fold (DN-FoldS), which showed the comparable results at the level of family and superfamily, compared to ensemble DN-Fold. Finally, we extended the binary classification problem of fold recognition to real-value regression task, which also show a promising performance. DN-Fold is freely available through a web server at http://iris.rnet.missouri.edu/dnfold.

  4. Improving Protein Fold Recognition by Deep Learning Networks

    NASA Astrophysics Data System (ADS)

    Jo, Taeho; Hou, Jie; Eickholt, Jesse; Cheng, Jianlin

    2015-12-01

    For accurate recognition of protein folds, a deep learning network method (DN-Fold) was developed to predict if a given query-template protein pair belongs to the same structural fold. The input used stemmed from the protein sequence and structural features extracted from the protein pair. We evaluated the performance of DN-Fold along with 18 different methods on Lindahl’s benchmark dataset and on a large benchmark set extracted from SCOP 1.75 consisting of about one million protein pairs, at three different levels of fold recognition (i.e., protein family, superfamily, and fold) depending on the evolutionary distance between protein sequences. The correct recognition rate of ensembled DN-Fold for Top 1 predictions is 84.5%, 61.5%, and 33.6% and for Top 5 is 91.2%, 76.5%, and 60.7% at family, superfamily, and fold levels, respectively. We also evaluated the performance of single DN-Fold (DN-FoldS), which showed the comparable results at the level of family and superfamily, compared to ensemble DN-Fold. Finally, we extended the binary classification problem of fold recognition to real-value regression task, which also show a promising performance. DN-Fold is freely available through a web server at http://iris.rnet.missouri.edu/dnfold.

  5. Improving Protein Fold Recognition by Deep Learning Networks.

    PubMed

    Jo, Taeho; Hou, Jie; Eickholt, Jesse; Cheng, Jianlin

    2015-01-01

    For accurate recognition of protein folds, a deep learning network method (DN-Fold) was developed to predict if a given query-template protein pair belongs to the same structural fold. The input used stemmed from the protein sequence and structural features extracted from the protein pair. We evaluated the performance of DN-Fold along with 18 different methods on Lindahl's benchmark dataset and on a large benchmark set extracted from SCOP 1.75 consisting of about one million protein pairs, at three different levels of fold recognition (i.e., protein family, superfamily, and fold) depending on the evolutionary distance between protein sequences. The correct recognition rate of ensembled DN-Fold for Top 1 predictions is 84.5%, 61.5%, and 33.6% and for Top 5 is 91.2%, 76.5%, and 60.7% at family, superfamily, and fold levels, respectively. We also evaluated the performance of single DN-Fold (DN-FoldS), which showed the comparable results at the level of family and superfamily, compared to ensemble DN-Fold. Finally, we extended the binary classification problem of fold recognition to real-value regression task, which also show a promising performance. DN-Fold is freely available through a web server at http://iris.rnet.missouri.edu/dnfold. PMID:26634993

  6. Coupled Folding and Specific Binding: Fishing for Amphiphilicity

    PubMed Central

    Jain, Vikas P.; Tu, Raymond S.

    2011-01-01

    Proteins are uniquely capable of identifying targets with unparalleled selectivity, but, in addition to the precision of the binding phenomenon, nature has the ability to find its targets exceptionally quickly. Transcription factors for instance can bind to a specific sequence of nucleic acids from a soup of similar, but not identical DNA strands, on a timescale of seconds. This is only possible with the enhanced kinetics provided for by a natively disordered structure, where protein folding and binding are cooperative processes. The secondary structures of many proteins are disordered under physiological conditions. Subsequently, the disordered structures fold into ordered structures only when they bind to their specific targets. Induced folding of the protein has two key biological advantages. First, flexible unstructured domains can result in an intrinsic plasticity that allows them to accommodate targets of various size and shape. And, second, the dynamics of this folding process can result in enhanced binding kinetics. Several groups have hypothesized the acceleration of binding kinetics is due to induced folding where a “fly-casting” effect has been shown to break the diffusion-limited rate of binding. This review describes experimental results in rationally designed peptide systems where the folding is coupled to amphiphilicity and biomolecular activity. PMID:21673899

  7. Exploring one-state downhill protein folding in single molecules

    PubMed Central

    Liu, Jianwei; Campos, Luis A.; Cerminara, Michele; Wang, Xiang; Ramanathan, Ravishankar; English, Douglas S.; Muñoz, Victor

    2012-01-01

    A one-state downhill protein folding process is barrierless at all conditions, resulting in gradual melting of native structure that permits resolving folding mechanisms step-by-step at atomic resolution. Experimental studies of one-state downhill folding have typically focused on the thermal denaturation of proteins that fold near the speed limit (ca. 106 s-1) at their unfolding temperature, thus being several orders of magnitude too fast for current single-molecule methods, such as single-molecule FRET. An important open question is whether one-state downhill folding kinetics can be slowed down to make them accessible to single-molecule approaches without turning the protein into a conventional activated folder. Here we address this question on the small helical protein BBL, a paradigm of one-state downhill thermal (un)folding. We decreased 200-fold the BBL folding-unfolding rate by combining chemical denaturation and low temperature, and carried out free-diffusion single-molecule FRET experiments with 50-μs resolution and maximal photoprotection using a recently developed Trolox-cysteamine cocktail. These experiments revealed a single conformational ensemble at all denaturing conditions. The chemical unfolding of BBL was then manifested by the gradual change of this unique ensemble, which shifts from high to low FRET efficiency and becomes broader at increasing denaturant. Furthermore, using detailed quantitative analysis, we could rule out the possibility that the BBL single-molecule data are produced by partly overlapping folded and unfolded peaks. Thus, our results demonstrate the one-state downhill folding regime at the single-molecule level and highlight that this folding scenario is not necessarily associated with ultrafast kinetics. PMID:22184219

  8. How the genome folds

    NASA Astrophysics Data System (ADS)

    Lieberman Aiden, Erez

    2012-02-01

    I describe Hi-C, a novel technology for probing the three-dimensional architecture of whole genomes by coupling proximity-based ligation with massively parallel sequencing. Working with collaborators at the Broad Institute and UMass Medical School, we used Hi-C to construct spatial proximity maps of the human genome at a resolution of 1Mb. These maps confirm the presence of chromosome territories and the spatial proximity of small, gene-rich chromosomes. We identified an additional level of genome organization that is characterized by the spatial segregation of open and closed chromatin to form two genome-wide compartments. At the megabase scale, the chromatin conformation is consistent with a fractal globule, a knot-free conformation that enables maximally dense packing while preserving the ability to easily fold and unfold any genomic locus. The fractal globule is distinct from the more commonly used globular equilibrium model. Our results demonstrate the power of Hi-C to map the dynamic conformations of whole genomes.

  9. Improved Method of Design for Folding Inflatable Shells

    NASA Technical Reports Server (NTRS)

    Johnson, Christopher J.

    2009-01-01

    An improved method of designing complexly shaped inflatable shells to be assembled from gores was conceived for original application to the inflatable outer shell of a developmental habitable spacecraft module having a cylindrical mid-length section with toroidal end caps. The method is also applicable to inflatable shells of various shapes for terrestrial use. The method addresses problems associated with the assembly, folding, transport, and deployment of inflatable shells that may comprise multiple layers and have complex shapes that can include such doubly curved surfaces as toroids and spheres. One particularly difficult problem is that of mathematically defining fold lines on a gore pattern in a double- curvature region. Moreover, because the fold lines in a double-curvature region tend to be curved, there is a practical problem of how to implement the folds. Another problem is that of modifying the basic gore shapes and sizes for the various layers so that when they are folded as part of the integral structure, they do not mechanically interfere with each other at the fold lines. Heretofore, it has been a common practice to design an inflatable shell to be assembled in the deployed configuration, without regard for the need to fold it into compact form. Typically, the result has been that folding has been a difficult, time-consuming process resulting in a An improved method of designing complexly shaped inflatable shells to be assembled from gores was conceived for original application to the inflatable outer shell of a developmental habitable spacecraft module having a cylindrical mid-length section with toroidal end caps. The method is also applicable to inflatable shells of various shapes for terrestrial use. The method addresses problems associated with the assembly, folding, transport, and deployment of inflatable shells that may comprise multiple layers and have complex shapes that can include such doubly curved surfaces as toroids and spheres. One

  10. How the folding rates of two- and multistate proteins depend on the amino acid properties.

    PubMed

    Huang, Jitao T; Huang, Wei; Huang, Shanran R; Li, Xin

    2014-10-01

    Proteins fold by either two-state or multistate kinetic mechanism. We observe that amino acids play different roles in different mechanism. Many residues that are easy to form regular secondary structures (α helices, β sheets and turns) can promote the two-state folding reactions of small proteins. Most of hydrophilic residues can speed up the multistate folding reactions of large proteins. Folding rates of large proteins are equally responsive to the flexibility of partial amino acids. Other properties of amino acids (including volume, polarity, accessible surface, exposure degree, isoelectric point, and phase transfer energy) have contributed little to folding kinetics of the proteins. Cysteine is a special residue, it triggers two-state folding reaction and but inhibits multistate folding reaction. These findings not only provide a new insight into protein structure prediction, but also could be used to direct the point mutations that can change folding rate.

  11. Slow and Bimolecular Folding of a De Novo Designed Monomeric Protein DS119

    PubMed Central

    Zhu, Cheng; Dai, Ziwei; Liang, Huanhuan; Zhang, Tao; Gai, Feng; Lai, Luhua

    2013-01-01

    De novo protein design offers a unique means to test and advance our understanding of how proteins fold. However, most current design methods are native structure eccentric and folding kinetics has rarely been considered in the design process. Here, we show that a de novo designed mini-protein DS119, which folds into a βαβ structure, exhibits unusually slow and concentration-dependent folding kinetics. For example, the folding time for 50 μM of DS119 was estimated to be ∼2 s. Stopped-flow fluorescence resonance energy transfer experiments further suggested that its folding was likely facilitated by a transient dimerization process. Taken together, these results highlight the need for consideration of the entire folding energy landscape in de novo protein design and provide evidence suggesting nonnative interactions can play a key role in protein folding. PMID:24209859

  12. Protein folding in the ER.

    SciTech Connect

    Stevens, F. J.; Argon, Y.; Biosciences Division; Univ. of Chicago

    1999-10-01

    The endoplasmic reticulum (ER) is a major protein folding compartment for secreted, plasma membrane and organelle proteins. Each of these newly-synthesized polypeptides folds in a deterministic process, affected by the unique conditions that exist in the ER. An understanding of protein folding in the ER is a fundamental biomolecular challenge at two levels. The first level addresses how the amino acid sequence programs that polypeptide to efficiently arrive at a particular fold out of a multitude of alternatives, and how different sequences obtain similar folds. At the second level are the issues introduced by folding not in the cytosol, but in the ER, including the risk of aggregation in a molecularly crowded environment, accommodation of post-translational modifications and the compatibility with subsequent intracellular trafficking. This review discusses both the physicochemical and cell biological constraints of folding, which are the challenges that the ER molecular chaperones help overcome.

  13. Structural origin of slow diffusion in protein folding.

    PubMed

    Chung, Hoi Sung; Piana-Agostinetti, Stefano; Shaw, David E; Eaton, William A

    2015-09-25

    Experimental, theoretical, and computational studies of small proteins suggest that interresidue contacts not present in the folded structure play little or no role in the self-assembly mechanism. Non-native contacts can, however, influence folding kinetics by introducing additional local minima that slow diffusion over the global free-energy barrier between folded and unfolded states. Here, we combine single-molecule fluorescence with all-atom molecular dynamics simulations to discover the structural origin for the slow diffusion that markedly decreases the folding rate for a designed α-helical protein. Our experimental determination of transition path times and our analysis of the simulations point to non-native salt bridges between helices as the source, which provides a quantitative glimpse of how specific intramolecular interactions influence protein folding rates by altering dynamics and not activation free energies.

  14. Rise of the Helix from a Collapsed Globule during the Folding of Monellin.

    PubMed

    Goluguri, Rama Reddy; Udgaonkar, Jayant B

    2015-09-01

    Early kinetic intermediates observed during the folding of many proteins are invariably compact and appear to possess some secondary structure. Consequently, it has been difficult to understand whether compaction drives secondary structure formation or secondary structure formation facilitates compaction during folding. In this study of the folding of single-chain monellin, it is shown that a kinetic molten globule (MG) is populated at 2 ms of folding. Far-UV circular dichroism (CD) measurements show that the kinetic MG is devoid of any helical structure even under the most stabilizing folding conditions. Multisite fluorescence resonance energy transfer (FRET) measurements show that the kinetic MG is compact with different segments having contracted to different extents. It is shown that the sequence segment that goes on to form the sole helix in the native protein is fully collapsed in the kinetic MG. This segment expands to accommodate the helix as the kinetic MG folds further to the native state, while other segments of the protein contract. Helix formation starting from the kinetic MG is shown to occur in multiple kinetic steps, whether measured by far-UV CD or by FRET. PMID:26258844

  15. Communication Between RNA Folding Domains Revealed by Folding of Circularly Permuted Ribozymes

    SciTech Connect

    Lease,R.; Adilakshmi, T.; Heilman-Miller, S.; Woodson, S.

    2007-01-01

    To study the role of sequence and topology in RNA folding, we determined the kinetic folding pathways of two circularly permuted variants of the Tetrahymena group I ribozyme, using time-resolved hydroxyl radical footprinting. Circular permutation changes the distance between interacting residues in the primary sequence, without changing the native structure of the RNA. In the natural ribozyme, tertiary interactions in the P4-P6 domain form in 1 s, while interactions in the P3-P9 form in 1-3 min at 42 C. Permutation of the 5' end to G111 in the P4 helix allowed the stable P4-P6 domain to fold in 200 ms at 30 C, five times faster than in the wild-type RNA, while the other domains folded five times more slowly (5-8 min). By contrast, circular permutation of the 5' end to G303 in J8/7 decreased the folding rate of the P4-P6 domain. In this permuted RNA, regions joining P2, P3 and P4 were protected in 500 ms, while the P3-P9 domain was 60-80% folded within 30 s. RNase T1 digestion and FMN photocleavage showed that circular permutation of the RNA sequence alters the initial ensemble of secondary structures, thereby changing the tertiary folding pathways. Our results show that the natural 5'-to-3' order of the structural domains in group I ribozymes optimizes structural communication between tertiary domains and promotes self-assembly of the catalytic center.

  16. Graphene folding on flat substrates

    SciTech Connect

    Chen, Xiaoming; Zhao, Yadong; Ke, Changhong; Zhang, Liuyang; Wang, Xianqiao

    2014-10-28

    We present a combined experimental-theoretical study of graphene folding on flat substrates. The structure and deformation of the folded graphene sheet are experimentally characterized by atomic force microscopy. The local graphene folding behaviors are interpreted based on nonlinear continuum mechanics modeling and molecular dynamics simulations. Our study on self-folding of a trilayer graphene sheet reports a bending stiffness of about 6.57 eV, which is about four times the reported values for monolayer graphene. Our results reveal that an intriguing free sliding phenomenon occurs at the interlayer van der Waals interfaces during the graphene folding process. This work demonstrates that it is a plausible venue to quantify the bending stiffness of graphene based on its self-folding conformation on flat substrates. The findings reported in this work are useful to a better understanding of the mechanical properties of graphene and in the pursuit of its applications.

  17. Abstract folding space analysis based on helices

    PubMed Central

    Huang, Jiabin; Backofen, Rolf; Voß, Björn

    2012-01-01

    RNA has many pivotal functions especially in the regulation of gene expression by ncRNAs. Identification of their structure is an important requirement for understanding their function. Structure prediction alone is often insufficient for this task, due to algorithmic problems, parameter inaccuracies, and biological peculiarities. Among the latter, there are base modifications, cotranscriptional folding leading to folding traps, and conformational switching as in the case of riboswitches. All these require more in-depth analysis of the folding space. The major drawback, which all methods have to cope with, is the exponential growth of the folding space. Therefore, methods are often limited in the sequence length they can analyze, or they make use of heuristics, sampling, or abstraction. Our approach adopts the abstraction strategy and remedies some problems of existing methods. We introduce a position-specific abstraction based on helices that we term helix index shapes, or hishapes for short. Utilizing a dynamic programming framework, we have implemented this abstraction in the program RNAHeliCes. Furthermore, we developed two hishape-based methods, one for energy barrier estimation, called HiPath, and one for abstract structure comparison, termed HiTed. We demonstrate the superior performance of HiPath compared to other existing methods and the competitive accuracy of HiTed. RNAHeliCes, together with HiPath and HiTed, are available for download at http://www.cyanolab.de/software/RNAHeliCes.htm. PMID:23104999

  18. Physical experiments of transpressional folding

    NASA Astrophysics Data System (ADS)

    Tikoff, Basil; Peterson, Karl

    1998-06-01

    In order to understand the process of folding in obliquely convergent settings, we formed folds within a shear box capable of creating homogeneous transpressional deformations. Folds were created in a single layer of stiff mixed plasticine and silicone that overlay a Newtonian silicone, for a variety of plate convergence angles. As small amplitude folds became visible, they were parallel to the long axis of the horizontal finite strain ellipse. With increasing deformation, the fold hinges rotated parallel with the long axis of the horizontal finite strain ellipse for all angles of convergence. This parallelism indicates that fold hinges, once formed, rotate with the horizontal strain ellipse rather than as material lines. The experiments highlight several interesting effects of transpression dynamics. The fold hinges initiate parallel to either ṡ1 or ṡ2 and are parallel to either S1 or S2 with increasing deformation. Neither infinitesimal strain (stress) nor finite strain is resolvable solely from fold geometry. Further, the net amount of contraction determined by folding across the zone was overestimated in all cases except pure contraction. This effect is obvious for the case of wrenching, where folding implies that the zone contracts if elongation parallel to the fold hinge is not considered. Therefore, attempts to balance cross-sections in transpressional zones will tend to overestimate contraction unless the wrench component of deformation is addressed. This result is validated by applying the modeling results in folding in central California adjacent to the San Andreas fault, where cross-section balancing indicates higher amounts of contraction than predicted by plate motion.

  19. COS NUV MAMA Fold Distribution

    NASA Astrophysics Data System (ADS)

    Wheeler, Thomas

    2010-09-01

    The performance of the MAMA microchannel plate can be monitored using a MAMA fold analysis procedure. The fold analysis provides a measurement of the distribution of charge cloud sizes incident upon the anode giving some measure of changes in the pulse-height distribution of the MCP and, therefore, MCP gain. This proposal executes the same steps as the COS MAMA Fold Analysis {11891} during Cycle 17.

  20. COS NUV MAMA Fold Distribution

    NASA Astrophysics Data System (ADS)

    Wheeler, Thomas

    2012-10-01

    The performance of the MAMA microchannel plate can be monitored using a MAMA fold analysis procedure. The fold analysis provides a measurement of the distribution of charge cloud sizes incident upon the anode giving some measure of changes in the pulse-height distribution of the MCP and, therefore, MCP gain. This proposal executes the same steps as the COS MAMA Fold Analysis {12723} during Cycle 19.

  1. WW Domain Folding Complexity Revealed by Infrared Spectroscopy

    PubMed Central

    2015-01-01

    Although the intrinsic tryptophan fluorescence of proteins offers a convenient probe of protein folding, interpretation of the fluorescence spectrum is often difficult because it is sensitive to both global and local changes. Infrared (IR) spectroscopy offers a complementary measure of structural changes involved in protein folding, because it probes changes in the secondary structure of the protein backbone. Here we demonstrate the advantages of using multiple probes, infrared and fluorescence spectroscopy, to study the folding of the FBP28 WW domain. Laser-induced temperature jumps coupled with fluorescence or infrared spectroscopy have been used to probe changes in the peptide backbone on the submillisecond time scale. The relaxation dynamics of the β-sheets and β-turn were measured independently by probing the corresponding IR bands assigned in the amide I region. Using these wavelength-dependent measurements, we observe three kinetics phases, with the fastest process corresponding to the relaxation kinetics of the turns. In contrast, fluorescence measurements of the wild-type WW domain and tryptophan mutants exhibit single-exponential kinetics with a lifetime that corresponds to the slowest phase observed by infrared spectroscopy. Mutant sequences provide evidence of an intermediate dry molten globule state. The slowest step in the folding of this WW domain is the tight packing of the side chains in the transition from the dry molten globule intermediate to the native structure. This study demonstrates that using multiple complementary probes enhances the interpretation of protein folding dynamics. PMID:25121968

  2. WW domain folding complexity revealed by infrared spectroscopy.

    PubMed

    Davis, Caitlin M; Dyer, R Brian

    2014-09-01

    Although the intrinsic tryptophan fluorescence of proteins offers a convenient probe of protein folding, interpretation of the fluorescence spectrum is often difficult because it is sensitive to both global and local changes. Infrared (IR) spectroscopy offers a complementary measure of structural changes involved in protein folding, because it probes changes in the secondary structure of the protein backbone. Here we demonstrate the advantages of using multiple probes, infrared and fluorescence spectroscopy, to study the folding of the FBP28 WW domain. Laser-induced temperature jumps coupled with fluorescence or infrared spectroscopy have been used to probe changes in the peptide backbone on the submillisecond time scale. The relaxation dynamics of the β-sheets and β-turn were measured independently by probing the corresponding IR bands assigned in the amide I region. Using these wavelength-dependent measurements, we observe three kinetics phases, with the fastest process corresponding to the relaxation kinetics of the turns. In contrast, fluorescence measurements of the wild-type WW domain and tryptophan mutants exhibit single-exponential kinetics with a lifetime that corresponds to the slowest phase observed by infrared spectroscopy. Mutant sequences provide evidence of an intermediate dry molten globule state. The slowest step in the folding of this WW domain is the tight packing of the side chains in the transition from the dry molten globule intermediate to the native structure. This study demonstrates that using multiple complementary probes enhances the interpretation of protein folding dynamics. PMID:25121968

  3. Compact intermediates in RNA folding

    SciTech Connect

    Woodson, S.A.

    2011-12-14

    Large noncoding RNAs fold into their biologically functional structures via compact yet disordered intermediates, which couple the stable secondary structure of the RNA with the emerging tertiary fold. The specificity of the collapse transition, which coincides with the assembly of helical domains, depends on RNA sequence and counterions. It determines the specificity of the folding pathways and the magnitude of the free energy barriers to the ensuing search for the native conformation. By coupling helix assembly with nascent tertiary interactions, compact folding intermediates in RNA also play a crucial role in ligand binding and RNA-protein recognition.

  4. Folding superfunnel to describe cooperative folding of interacting proteins.

    PubMed

    Smeller, László

    2016-07-01

    This paper proposes a generalization of the well-known folding funnel concept of proteins. In the funnel model the polypeptide chain is treated as an individual object not interacting with other proteins. Since biological systems are considerably crowded, protein-protein interaction is a fundamental feature during the life cycle of proteins. The folding superfunnel proposed here describes the folding process of interacting proteins in various situations. The first example discussed is the folding of the freshly synthesized protein with the aid of chaperones. Another important aspect of protein-protein interactions is the folding of the recently characterized intrinsically disordered proteins, where binding to target proteins plays a crucial role in the completion of the folding process. The third scenario where the folding superfunnel is used is the formation of aggregates from destabilized proteins, which is an important factor in case of several conformational diseases. The folding superfunnel constructed here with the minimal assumption about the interaction potential explains all three cases mentioned above. Proteins 2016; 84:1009-1016. © 2016 Wiley Periodicals, Inc.

  5. Folding and unfolding single RNA molecules under tension

    PubMed Central

    Woodside, Michael T; García-García, Cuauhtémoc; Block, Steven M

    2010-01-01

    Single-molecule force spectroscopy constitutes a powerful method for probing RNA folding: it allows the kinetic, energetic, and structural properties of intermediate and transition states to be determined quantitatively, yielding new insights into folding pathways and energy landscapes. Recent advances in experimental and theoretical methods, including fluctuation theorems, kinetic theories, novel force clamps, and ultrastable instruments, have opened new avenues for study. These tools have been used to probe folding in simple model systems, for example, RNA and DNA hairpins. Knowledge gained from such systems is helping to build our understanding of more complex RNA structures composed of multiple elements, as well as how nucleic acids interact with proteins involved in key cellular activities, such as transcription and translation. PMID:18786653

  6. Characterization of a folding intermediate from HIV-1 ribonuclease H.

    PubMed Central

    Kern, G.; Handel, T.; Marqusee, S.

    1998-01-01

    The RNase H domain from HIV-1 (HIV RNase H) encodes an essential retroviral activity. Refolding of the isolated HIV RNase H domain shows a kinetic intermediate detectable by stopped-flow far UV circular dichroism and pulse-labeling H/D exchange. In this intermediate, strands 1, 4, and 5 as well as helices A and D appear to be structured. Compared to its homolog from Escherichia coli, the rate limiting step in refolding of HIV RNase H appears closer to the native state. We have modeled this kinetic intermediate using a C-terminal deletion fragment lacking helix E. Like the kinetic intermediate, this variant folds rapidly and shows a decrease in stability. We propose that inhibition of the docking of helix E to this folding intermediate may present a novel strategy for anti HIV-1 therapy. PMID:9792104

  7. Structural Bridges through Fold Space

    PubMed Central

    Edwards, Hannah; Deane, Charlotte M.

    2015-01-01

    Several protein structure classification schemes exist that partition the protein universe into structural units called folds. Yet these schemes do not discuss how these units sit relative to each other in a global structure space. In this paper we construct networks that describe such global relationships between folds in the form of structural bridges. We generate these networks using four different structural alignment methods across multiple score thresholds. The networks constructed using the different methods remain a similar distance apart regardless of the probability threshold defining a structural bridge. This suggests that at least some structural bridges are method specific and that any attempt to build a picture of structural space should not be reliant on a single structural superposition method. Despite these differences all representations agree on an organisation of fold space into five principal community structures: all-α, all-β sandwiches, all-β barrels, α/β and α + β. We project estimated fold ages onto the networks and find that not only are the pairings of unconnected folds associated with higher age differences than bridged folds, but this difference increases with the number of networks displaying an edge. We also examine different centrality measures for folds within the networks and how these relate to fold age. While these measures interpret the central core of fold space in varied ways they all identify the disposition of ancestral folds to fall within this core and that of the more recently evolved structures to provide the peripheral landscape. These findings suggest that evolutionary information is encoded along these structural bridges. Finally, we identify four highly central pivotal folds representing dominant topological features which act as key attractors within our landscapes. PMID:26372166

  8. Structural Bridges through Fold Space.

    PubMed

    Edwards, Hannah; Deane, Charlotte M

    2015-09-01

    Several protein structure classification schemes exist that partition the protein universe into structural units called folds. Yet these schemes do not discuss how these units sit relative to each other in a global structure space. In this paper we construct networks that describe such global relationships between folds in the form of structural bridges. We generate these networks using four different structural alignment methods across multiple score thresholds. The networks constructed using the different methods remain a similar distance apart regardless of the probability threshold defining a structural bridge. This suggests that at least some structural bridges are method specific and that any attempt to build a picture of structural space should not be reliant on a single structural superposition method. Despite these differences all representations agree on an organisation of fold space into five principal community structures: all-α, all-β sandwiches, all-β barrels, α/β and α + β. We project estimated fold ages onto the networks and find that not only are the pairings of unconnected folds associated with higher age differences than bridged folds, but this difference increases with the number of networks displaying an edge. We also examine different centrality measures for folds within the networks and how these relate to fold age. While these measures interpret the central core of fold space in varied ways they all identify the disposition of ancestral folds to fall within this core and that of the more recently evolved structures to provide the peripheral landscape. These findings suggest that evolutionary information is encoded along these structural bridges. Finally, we identify four highly central pivotal folds representing dominant topological features which act as key attractors within our landscapes.

  9. Folding λ-Repressor at Its Speed Limit

    PubMed Central

    Yang, Wei Yuan; Gruebele, Martin

    2004-01-01

    We show that the five-helix bundle λ6–85 can be engineered and solvent-tuned to make the transition from activated two-state folding to downhill folding. The transition manifests itself as the appearance of additional dynamics faster than the activated kinetics, followed by the disappearance of the activated kinetics when the bias toward the native state is increased. Our fastest value of 1 μs for the “speed” limit of λ6–85 is measured at low concentrations of a denaturant that smoothes the free-energy surface. Complete disappearance of the activated phase is obtained in stabilizing glucose buffer. Langevin dynamics on a rough free-energy surface with variable bias toward the native state provides a robust and quantitative description of the transition from activated to downhill folding. Based on our simulation, we estimate the residual energetic frustration of λ6–85 to be δ2 G ≈ 0.64 k2T2. We show that λ6–86, as well as very fast folding proteins or folding intermediates estimated to lie near the speed limit, provide a better rate-topology correlation than proteins with larger energetic frustration. A limit of β ≥ 0.7 on any stretching of λ6–85 barrier-free dynamics suggests that a low-dimensional free-energy surface is sufficient to describe folding. PMID:15240492

  10. Novel sequences propel familiar folds.

    PubMed

    Jawad, Zahra; Paoli, Massimo

    2002-04-01

    Recent structure determinations have made new additions to a set of strikingly different sequences that give rise to the same topology. Proteins with a beta propeller fold are characterized by extreme sequence diversity despite the similarity in their three-dimensional structures. Several fold predictions, based in part on sequence repeats thought to match modular beta sheets, have been proved correct.

  11. COS NUV MAMA Fold Distribution

    NASA Astrophysics Data System (ADS)

    Wheeler, Thomas

    2013-10-01

    The performance of the MAMA microchannel plate can be monitored using a MAMA fold analysis procedure. The fold analysis provides a measurement of the distribution of charge cloud sizes incident upon the anode giving some measure of changes in the pulse-height distribution of the MCP and, therefore, MCP gain. This proposal executes the same steps as Cycle 20 proposal 13128.

  12. When Do Semi-crystalline Polymer Fold during Crystallization?

    NASA Astrophysics Data System (ADS)

    Miyoshi, Toshikazu; Hong, You-Lee

    2014-03-01

    Semi-crystalline polymers are crystallized as folded chains in thin lamellae of ca. 5-20 nm from random coils in the melt and solution states. Lauritzen-Hoffman theory implied the crystallization process is dominated by sequential stem deposition on the growth front. Conversely, Allegra proposed a bundle model in which aggregates of 10-20 stems are produced by folding in the pre-stage of crystallization. The pre-folded chains are kinetically deposited on the growth front and thus determine the morphology at different crystallization temperatures. The folded chains preserve their own chain-folding directions, numbers, and fractions as a function of concentrations and supercooling, which would provide detailed chain-folding mechanism. We recently developed a new strategy using 13C-13C double-quantum NMR to investigate chain-trajectory of 13C selectively labeled polymer in bulk crystals. Here, we report how re-entrance sites, fraction, and number of folded chains of isotactic poly(1-butene) in form III single crystals depends on supercooling conditions and solvent effects. On the basis of molecular level structures, we will discuss about chain-folding process of iPB1 in dilute solutions. DMR-1105829.

  13. Folding and escape of nascent proteins at ribosomal exit tunnel

    NASA Astrophysics Data System (ADS)

    Bui, Phuong Thuy; Hoang, Trinh Xuan

    2016-03-01

    We investigate the interplay between post-translational folding and escape of two small single-domain proteins at the ribosomal exit tunnel by using Langevin dynamics with coarse-grained models. It is shown that at temperatures lower or near the temperature of the fastest folding, folding proceeds concomitantly with the escape process, resulting in vectorial folding and enhancement of foldability of nascent proteins. The concomitance between the two processes, however, deteriorates as temperature increases. Our folding simulations as well as free energy calculation by using umbrella sampling show that, at low temperatures, folding at the tunnel follows one or two specific pathways without kinetic traps. It is shown that the escape time can be mapped to a one-dimensional diffusion model with two different regimes for temperatures above and below the folding transition temperature. Attractive interactions between amino acids and attractive sites on the tunnel wall lead to a free energy barrier along the escape route of the protein. It is suggested that this barrier slows down the escape process and consequently promotes correct folding of the released nascent protein.

  14. Activation Barrier-Limited Folding and Conformational Sampling of a Dynamic Protein Domain.

    PubMed

    Dogan, Jakob; Toto, Angelo; Andersson, Eva; Gianni, Stefano; Jemth, Per

    2016-09-20

    Folding reaction mechanisms of globular protein domains have been extensively studied by both experiment and simulation and found to be highly concerted chemical reactions in which numerous noncovalent bonds form in an apparent two-state fashion. However, less is known regarding intrinsically disordered proteins because their folding can usually be studied only in conjunction with binding to a ligand. We have investigated by kinetics the folding mechanism of such a disordered protein domain, the nuclear coactivator-binding domain (NCBD) from CREB-binding protein. While a previous computational study suggested that NCBD folds without an activation free energy barrier, our experimental data demonstrate that NCBD, despite its highly dynamic structure, displays relatively slow folding (∼10 ms at 277 K) consistent with a barrier-limited process. Furthermore, the folding kinetics corroborate previous nuclear magnetic resonance data showing that NCBD exists in two folded conformations and one more denatured conformation at equilibrium and, thus, that the folding mechanism is a three-state mechanism. The refolding kinetics is limited by unfolding of the less populated folded conformation, suggesting that the major route for interconversion between the two folded states is via the denatured state. Because the two folded conformations have been suggested to bind distinct ligands, our results have mechanistic implications for conformational sampling in protein-protein interactions. PMID:27542287

  15. Periodic folding of viscous sheets

    NASA Astrophysics Data System (ADS)

    Ribe, Neil M.

    2003-09-01

    The periodic folding of a sheet of viscous fluid falling upon a rigid surface is a common fluid mechanical instability that occurs in contexts ranging from food processing to geophysics. Asymptotic thin-layer equations for the combined stretching-bending deformation of a two-dimensional sheet are solved numerically to determine the folding frequency as a function of the sheet’s initial thickness, the pouring speed, the height of fall, and the fluid properties. As the buoyancy increases, the system bifurcates from “forced” folding driven kinematically by fluid extrusion to “free” folding in which viscous resistance to bending is balanced by buoyancy. The systematics of the numerically predicted folding frequency are in good agreement with laboratory experiments.

  16. Multidimensional theory of protein folding

    NASA Astrophysics Data System (ADS)

    Itoh, Kazuhito; Sasai, Masaki

    2009-04-01

    Theory of multidimensional representation of free energy surface of protein folding is developed by adopting structural order parameters of multiple regions in protein as multiple coordinates. Various scenarios of folding are classified in terms of cooperativity within individual regions and interactions among multiple regions and thus obtained classification is used to analyze the folding process of several example proteins. Ribosomal protein S6, src-SH3 domain, CheY, barnase, and BBL domain are analyzed with the two-dimensional representation by using a structure-based Hamiltonian model. The extension to the higher dimensional representation leads to the finer description of the folding process. Barnase, NtrC, and an ankyrin repeat protein are examined with the three-dimensional representation. The multidimensional representation allows us to directly address questions on folding pathways, intermediates, and transition states.

  17. When fast is better: protein folding fundamentals and mechanisms from ultrafast approaches

    PubMed Central

    Muñoz, Victor; Cerminara, Michele

    2016-01-01

    Protein folding research stalled for decades because conventional experiments indicated that proteins fold slowly and in single strokes, whereas theory predicted a complex interplay between dynamics and energetics resulting in myriad microscopic pathways. Ultrafast kinetic methods turned the field upside down by providing the means to probe fundamental aspects of folding, test theoretical predictions and benchmark simulations. Accordingly, experimentalists could measure the timescales for all relevant folding motions, determine the folding speed limit and confirm that folding barriers are entropic bottlenecks. Moreover, a catalogue of proteins that fold extremely fast (microseconds) could be identified. Such fast-folding proteins cross shallow free energy barriers or fold downhill, and thus unfold with minimal co-operativity (gradually). A new generation of thermodynamic methods has exploited this property to map folding landscapes, interaction networks and mechanisms at nearly atomic resolution. In parallel, modern molecular dynamics simulations have finally reached the timescales required to watch fast-folding proteins fold and unfold in silico. All of these findings have buttressed the fundamentals of protein folding predicted by theory, and are now offering the first glimpses at the underlying mechanisms. Fast folding appears to also have functional implications as recent results connect downhill folding with intrinsically disordered proteins, their complex binding modes and ability to moonlight. These connections suggest that the coupling between downhill (un)folding and binding enables such protein domains to operate analogically as conformational rheostats. PMID:27574021

  18. When fast is better: protein folding fundamentals and mechanisms from ultrafast approaches.

    PubMed

    Muñoz, Victor; Cerminara, Michele

    2016-09-01

    Protein folding research stalled for decades because conventional experiments indicated that proteins fold slowly and in single strokes, whereas theory predicted a complex interplay between dynamics and energetics resulting in myriad microscopic pathways. Ultrafast kinetic methods turned the field upside down by providing the means to probe fundamental aspects of folding, test theoretical predictions and benchmark simulations. Accordingly, experimentalists could measure the timescales for all relevant folding motions, determine the folding speed limit and confirm that folding barriers are entropic bottlenecks. Moreover, a catalogue of proteins that fold extremely fast (microseconds) could be identified. Such fast-folding proteins cross shallow free energy barriers or fold downhill, and thus unfold with minimal co-operativity (gradually). A new generation of thermodynamic methods has exploited this property to map folding landscapes, interaction networks and mechanisms at nearly atomic resolution. In parallel, modern molecular dynamics simulations have finally reached the timescales required to watch fast-folding proteins fold and unfold in silico All of these findings have buttressed the fundamentals of protein folding predicted by theory, and are now offering the first glimpses at the underlying mechanisms. Fast folding appears to also have functional implications as recent results connect downhill folding with intrinsically disordered proteins, their complex binding modes and ability to moonlight. These connections suggest that the coupling between downhill (un)folding and binding enables such protein domains to operate analogically as conformational rheostats. PMID:27574021

  19. Polymer Uncrossing and Knotting in Protein Folding, and Their Role in Minimal Folding Pathways

    PubMed Central

    Mohazab, Ali R.; Plotkin, Steven S.

    2013-01-01

    We introduce a method for calculating the extent to which chain non-crossing is important in the most efficient, optimal trajectories or pathways for a protein to fold. This involves recording all unphysical crossing events of a ghost chain, and calculating the minimal uncrossing cost that would have been required to avoid such events. A depth-first tree search algorithm is applied to find minimal transformations to fold , , , and knotted proteins. In all cases, the extra uncrossing/non-crossing distance is a small fraction of the total distance travelled by a ghost chain. Different structural classes may be distinguished by the amount of extra uncrossing distance, and the effectiveness of such discrimination is compared with other order parameters. It was seen that non-crossing distance over chain length provided the best discrimination between structural and kinetic classes. The scaling of non-crossing distance with chain length implies an inevitable crossover to entanglement-dominated folding mechanisms for sufficiently long chains. We further quantify the minimal folding pathways by collecting the sequence of uncrossing moves, which generally involve leg, loop, and elbow-like uncrossing moves, and rendering the collection of these moves over the unfolded ensemble as a multiple-transformation “alignment”. The consensus minimal pathway is constructed and shown schematically for representative cases of an , , and knotted protein. An overlap parameter is defined between pathways; we find that proteins have minimal overlap indicating diverse folding pathways, knotted proteins are highly constrained to follow a dominant pathway, and proteins are somewhere in between. Thus we have shown how topological chain constraints can induce dominant pathway mechanisms in protein folding. PMID:23365638

  20. Distinct folding pathways of two homologous disulfide proteins: bovine pancreatic trypsin inhibitor and tick anticoagulant peptide.

    PubMed

    Chang, Jui-Yoa

    2011-01-01

    The folding pathways of disulfide proteins vary substantially (Arolas et al., Trends Biochem Sci 31: 292-301, 2006). The diversity is mainly manifested by (a) the extent of heterogeneity of folding intermediates, (b) the extent of presence of native-like intermediates, and (c) the variation of folding kinetics. Even among structurally similar proteins, the difference can be enormous. This is demonstrated in this concise review with two structurally homologous kunitz-type protease inhibitors, bovine pancreatic trypsin inhibitor and tick anticoagulant peptide, as well as a group of cystine knot proteins. The diversity of their folding mechanisms is illustrated with two different folding techniques: (a) the conventional method of disulfide oxidation (oxidative folding), and (b) the novel method of disulfide scrambling (Chang, J Biol Chem 277: 120-126, 2002). This review also highlights the convergence of folding models concluded form the conventional conformational folding and those obtained by oxidative folding.

  1. UFO (UnFold Operator) computer program abstract

    SciTech Connect

    Kissel, L.; Biggs, F.

    1982-11-01

    UFO (UnFold Operator) is an interactive user-oriented computer program designed to solve a wide range of problems commonly encountered in physical measurements. This document provides a summary of the capabilities of version 3A of UFO.

  2. Microfluidic Mixers for Studying Protein Folding

    PubMed Central

    Waldauer, Steven A.; Wu, Ling; Yao, Shuhuai; Bakajin, Olgica; Lapidus, Lisa J.

    2012-01-01

    The process by which a protein folds into its native conformation is highly relevant to biology and human health yet still poorly understood. One reason for this is that folding takes place over a wide range of timescales, from nanoseconds to seconds or longer, depending on the protein1. Conventional stopped-flow mixers have allowed measurement of folding kinetics starting at about 1 ms. We have recently developed a microfluidic mixer that dilutes denaturant ~100-fold in ~8 μs2. Unlike a stopped-flow mixer, this mixer operates in the laminar flow regime in which turbulence does not occur. The absence of turbulence allows precise numeric simulation of all flows within the mixer with excellent agreement to experiment3-4. Laminar flow is achieved for Reynolds numbers Re ≤100. For aqueous solutions, this requires micron scale geometries. We use a hard substrate, such as silicon or fused silica, to make channels 5-10 μm wide and 10 μm deep (See Figure 1). The smallest dimensions, at the entrance to the mixing region, are on the order of 1 μm in size. The chip is sealed with a thin glass or fused silica coverslip for optical access. Typical total linear flow rates are ~1 m/s, yielding Re~10, but the protein consumption is only ~0.5 nL/s or 1.8 μL/hr. Protein concentration depends on the detection method: For tryptophan fluorescence the typical concentration is 100 μM (for 1 Trp/protein) and for FRET the typical concentration is ~100 nM. The folding process is initiated by rapid dilution of denaturant from 6 M to 0.06 M guanidine hydrochloride. The protein in high denaturant flows down a central channel and is met on either side at the mixing region by buffer without denaturant moving ~100 times faster (see Figure 2). This geometry causes rapid constriction of the protein flow into a narrow jet ~100 nm wide. Diffusion of the light denaturant molecules is very rapid, while diffusion of the heavy protein molecules is much slower, diffusing less than 1 μm in 1 ms

  3. Accurate prediction of cellular co-translational folding indicates proteins can switch from post- to co-translational folding

    PubMed Central

    Nissley, Daniel A.; Sharma, Ajeet K.; Ahmed, Nabeel; Friedrich, Ulrike A.; Kramer, Günter; Bukau, Bernd; O'Brien, Edward P.

    2016-01-01

    The rates at which domains fold and codons are translated are important factors in determining whether a nascent protein will co-translationally fold and function or misfold and malfunction. Here we develop a chemical kinetic model that calculates a protein domain's co-translational folding curve during synthesis using only the domain's bulk folding and unfolding rates and codon translation rates. We show that this model accurately predicts the course of co-translational folding measured in vivo for four different protein molecules. We then make predictions for a number of different proteins in yeast and find that synonymous codon substitutions, which change translation-elongation rates, can switch some protein domains from folding post-translationally to folding co-translationally—a result consistent with previous experimental studies. Our approach explains essential features of co-translational folding curves and predicts how varying the translation rate at different codon positions along a transcript's coding sequence affects this self-assembly process. PMID:26887592

  4. Accurate prediction of cellular co-translational folding indicates proteins can switch from post- to co-translational folding.

    PubMed

    Nissley, Daniel A; Sharma, Ajeet K; Ahmed, Nabeel; Friedrich, Ulrike A; Kramer, Günter; Bukau, Bernd; O'Brien, Edward P

    2016-01-01

    The rates at which domains fold and codons are translated are important factors in determining whether a nascent protein will co-translationally fold and function or misfold and malfunction. Here we develop a chemical kinetic model that calculates a protein domain's co-translational folding curve during synthesis using only the domain's bulk folding and unfolding rates and codon translation rates. We show that this model accurately predicts the course of co-translational folding measured in vivo for four different protein molecules. We then make predictions for a number of different proteins in yeast and find that synonymous codon substitutions, which change translation-elongation rates, can switch some protein domains from folding post-translationally to folding co-translationally--a result consistent with previous experimental studies. Our approach explains essential features of co-translational folding curves and predicts how varying the translation rate at different codon positions along a transcript's coding sequence affects this self-assembly process. PMID:26887592

  5. Fast events in protein folding

    SciTech Connect

    Woodruff, W.; Callender, R.; Causgrove, T.; Dyer, R.; Williams, S.

    1996-04-01

    The primary objective of this work was to develop a molecular understanding of how proteins achieve their native three-dimensional (folded) structures. This requires the identification and characterization of intermediates in the protein folding process on all relevant timescales, from picoseconds to seconds. The short timescale events in protein folding have been entirely unknown. Prior to this work, state-of-the-art experimental approaches were limited to milliseconds or longer, when much of the folding process is already over. The gap between theory and experiment is enormous: current theoretical and computational methods cannot realistically model folding processes with lifetimes longer than one nanosecond. This unique approach to employ laser pump-probe techniques that combine novel methods of laser flash photolysis with time-resolved vibrational spectroscopic probes of protein transients. In this scheme, a short (picosecond to nanosecond) laser photolysis pulse was used to produce an instantaneous pH or temperature jump, thereby initiating a protein folding or unfolding reaction. Structure-specific, time-resolved vibrational probes were then used to identify and characterize protein folding intermediates.

  6. Coiling and Folding of Viscoelastic Jets

    NASA Astrophysics Data System (ADS)

    Majmudar, Trushant; Varagnat, Matthieu; McKinley, Gareth

    2007-11-01

    The study of fluid jets impacting on a flat surface has industrial applications in many areas, including processing of foods and consumer goods, bottle filling, and polymer melt processing. Previous studies have focused primarily on purely viscous, Newtonian fluids, which exhibit a number of different dynamical regimes including dripping, steady jetting, folding, and steady coiling. Here we add another dimension to the problem by focusing on mobile (low viscosity) viscoelastic fluids, with the study of two wormlike-micellar fluids, a cetylpyridinum-salicylic acid salt (CPyCl/NaSal) solution, and an industrially relevant shampoo base. We investigate the effects of viscosity and elasticity on the dynamics of axi-symmetric jets. The viscoelasticity of the fluids is systematically controlled by varying the concentration of salt counterions. Experimental methods include shear and extensional rheology measurements to characterize the fluids, and high-speed digital video imaging. In addition to the regimes observed in purely viscous systems, we also find a novel regime in which the elastic jet buckles and folds on itself, and alternates between coiling and folding behavior. We suggest phase diagrams and scaling laws for the coiling and folding frequencies through a systematic exploration of the experimental parameter space (height of fall, imposed flow rate, elasticity of the solution).

  7. Toward understanding driving forces in membrane protein folding.

    PubMed

    Hong, Heedeok

    2014-12-15

    α-Helical membrane proteins are largely composed of nonpolar residues that are embedded in the lipid bilayer. An enigma in the folding of membrane proteins is how a polypeptide chain can be condensed into the compact folded state in the environment where the hydrophobic effect cannot strongly drive molecular interactions. Probably other forces such as van der Waals packing, hydrogen bonding, and weakly polar interactions, which are regarded less important in the folding of water-soluble proteins, should emerge. However, it is not clearly understood how those individual forces operate and how they are balanced for stabilizing membrane proteins. Studying this problem is not a trivial task mainly because of the methodological challenges in controlling the reversible folding of membrane proteins in the lipid bilayer. Overcoming the hurdles, meaningful progress has been made in the field in the last few decades. This review will focus on recent studies tackling the problem of driving forces in membrane protein folding. PMID:25107533

  8. Complex Pathways in Folding of Protein G Explored by Simulation and Experiment

    PubMed Central

    Lapidus, Lisa J.; Acharya, Srabasti; Schwantes, Christian R.; Wu, Ling; Shukla, Diwakar; King, Michael; DeCamp, Stephen J.; Pande, Vijay S.

    2014-01-01

    The B1 domain of protein G has been a classic model system of folding for decades, the subject of numerous experimental and computational studies. Most of the experimental work has focused on whether the protein folds via an intermediate, but the evidence is mostly limited to relatively slow kinetic observations with a few structural probes. In this work we observe folding on the submillisecond timescale with microfluidic mixers using a variety of probes including tryptophan fluorescence, circular dichroism, and photochemical oxidation. We find that each probe yields different kinetics and compare these observations with a Markov State Model constructed from large-scale molecular dynamics simulations and find a complex network of states that yield different kinetics for different observables. We conclude that there are many folding pathways before the final folding step and that these paths do not have large free energy barriers. PMID:25140430

  9. Impact of structure space continuity on protein fold classification

    PubMed Central

    Xu, Jinrui; Zhang, Jianzhi

    2016-01-01

    Protein structure classification hierarchically clusters domain structures based on structure and/or sequence similarities and plays important roles in the study of protein structure-function relationship and protein evolution. Among many classifications, SCOP and CATH are widely viewed as the gold standards. Fold classification is of special interest because this is the lowest level of classification that does not depend on protein sequence similarity. The current fold classifications such as those in SCOP and CATH are controversial because they implicitly assume that folds are discrete islands in the structure space, whereas increasing evidence suggests significant similarities among folds and supports a continuous fold space. Although this problem is widely recognized, its impact on fold classification has not been quantitatively evaluated. Here we develop a likelihood method to classify a domain into the existing folds of CATH or SCOP using both query-fold structure similarities and within-fold structure heterogeneities. The new classification differs from the original classification for 3.4–12% of domains, depending on factors such as the structure similarity score and original classification scheme used. Because these factors differ for different biological purposes, our results indicate that the importance of considering structure space continuity in fold classification depends on the specific question asked. PMID:27006112

  10. Fog spontaneously folds mosquito wings

    NASA Astrophysics Data System (ADS)

    Dickerson, Andrew K.; Liu, Xing; Zhu, Ting; Hu, David L.

    2015-02-01

    The flexibility of insect wings confers aerodynamic benefits, but can also present a hazard if exposed to fog or dew. Fog can cause water to accumulate on wings, bending them into tight taco shapes and rendering them useless for flight. In this combined experimental and theoretical study, we use high-speed video to film the spontaneous folding of isolated mosquito wings due to the evaporation of a water drop. We predict shapes of the deformed wing using two-dimensional elastica theory, considering both surface tension and Laplace pressure. We also recommend fold-resistant geometries for the wings of flapping micro-aerial vehicles. Our work reveals the mechanism of insect wing folding and provides a framework for further study of capillarity-driven folding in both natural and biomimetic systems at small scales.

  11. Molecular dynamics studies of protein folding and aggregation

    NASA Astrophysics Data System (ADS)

    Ding, Feng

    This thesis applies molecular dynamics simulations and statistical mechanics to study: (i) protein folding; and (ii) protein aggregation. Most small proteins fold into their native states via a first-order-like phase transition with a major free energy barrier between the folded and unfolded states. A set of protein conformations corresponding to the free energy barrier, Delta G >> kBT, are the folding transition state ensemble (TSE). Due to their evasive nature, TSE conformations are hard to capture (probability ∝ exp(-DeltaG/k BT)) and characterize. A coarse-grained discrete molecular dynamics model with realistic steric constraints is constructed to reproduce the experimentally observed two-state folding thermodynamics. A kinetic approach is proposed to identify the folding TSE. A specific set of contacts, common to the TSE conformations, is identified as the folding nuclei which are necessary to be formed in order for the protein to fold. Interestingly, the amino acids at the site of the identified folding nuclei are highly conserved for homologous proteins sharing the same structures. Such conservation suggests that amino acids that are important for folding kinetics are under selective pressure to be preserved during the course of molecular evolution. In addition, studies of the conformations close to the transition states uncover the importance of topology in the construction of order parameter for protein folding transition. Misfolded proteins often form insoluble aggregates, amyloid fibrils, that deposit in the extracellular space and lead to a type of disease known as amyloidosis. Due to its insoluble and non-crystalline nature, the aggregation structure and, thus the aggregation mechanism, has yet to be uncovered. Discrete molecular dynamics studies reveal an aggregate structure with the same structural signatures as in experimental observations and show a nucleation aggregation scenario. The simulations also suggest a generic aggregation mechanism

  12. Protein folding by motion planning

    NASA Astrophysics Data System (ADS)

    Thomas, Shawna; Song, Guang; Amato, Nancy M.

    2005-12-01

    We investigate a novel approach for studying protein folding that has evolved from robotics motion planning techniques called probabilistic roadmap methods (PRMs). Our focus is to study issues related to the folding process, such as the formation of secondary and tertiary structures, assuming we know the native fold. A feature of our PRM-based framework is that the large sets of folding pathways in the roadmaps it produces, in just a few hours on a desktop PC, provide global information about the protein's energy landscape. This is an advantage over other simulation methods such as molecular dynamics or Monte Carlo methods which require more computation and produce only a single trajectory in each run. In our initial studies, we obtained encouraging results for several small proteins. In this paper, we investigate more sophisticated techniques for analyzing the folding pathways in our roadmaps. In addition to more formally revalidating our previous results, we present a case study showing that our technique captures known folding differences between the structurally similar proteins G and L. This research was supported in part by NSF CAREER Award CCR-9624315, NSF Grants ACI-9872126, EIA-9975018, EIA-0103742, EIA-9805823, ACR-0113971, CCR-0113974, EIA-9810937, EIA-0079874 and the Texas Higher Education Coordinating Board grant ATP-000512-0261-2001. ST was supported in part by an NSF Graduate Research Fellowship. GS was supported in part by an IBM PhD Fellowship.

  13. Electrochemistry of folded graphene edges.

    PubMed

    Ambrosi, Adriano; Bonanni, Alessandra; Pumera, Martin

    2011-05-01

    There is enormous interest in the investigation of electron transfer rates at the edges of graphene due to possible energy storage and sensing applications. While electrochemistry at the edges and the basal plane of graphene has been studied in the past, the new frontier is the electrochemistry of folded graphene edges. Here we describe the electrochemistry of folded graphene edges and compare it to that of open graphene edges. The materials were characterized in detail by high-resolution transmission electron microscopy, Raman spectroscopy, high-resolution X-ray photoelectron spectroscopy, electrochemical impedance spectroscopy and cyclic voltammetry. We found that the heterogeneous electron transfer rate is significantly lower on folded graphene edges compared to open edge sites for ferro/ferricyanide, and that electrochemical properties of open edges offer lower potential detection of biomarkers than the folded ones. It is apparent, therefore, that for sensing and biosensing applications the folded edges are less active than open edges, which should then be preferred for such applications. As folded edges are the product of thermal treatment of multilayer graphene, such thermal procedures should be avoided when fabricating graphene for electrochemical applications.

  14. The prosegment catalyzes native folding of Plasmodium falciparum plasmepsin II.

    PubMed

    Jaafar, Ahmad Haniff; Xiao, Huogen; Dee, Derek R; Bryksa, Brian C; Bhaumik, Prasenjit; Yada, Rickey Y

    2016-10-01

    Plasmepsin II is a malarial pepsin-like aspartic protease produced as a zymogen containing an N-terminal prosegment domain that is removed during activation. Despite structural similarities between active plasmepsin II and pepsin, their prosegments adopt different conformations in the respective zymogens. In contrast to pepsinogen, the proplasmepsin II prosegment is 80 residues longer, contains a transmembrane region and is non-essential for recombinant expression in an active form, thus calling into question the prosegment's precise function. The present study examines the role of the prosegment in the folding mechanism of plasmepsin II. Both a shorter (residues 77-124) and a longer (residues 65-124) prosegment catalyze plasmepsin II folding at rates more than four orders of magnitude faster compared to folding without prosegment. Native plasmepsin II is kinetically trapped and requires the prosegment both to catalyze folding and to shift the folding equilibrium towards the native conformation. Thus, despite low sequence identity and distinct zymogen conformations, the folding landscapes of plasmepsin II and pepsin, both with and without prosegment, are qualitatively identical. These results imply a conserved and unusual feature of the pepsin-like protease topology that necessitates prosegment-assisted folding. PMID:27378574

  15. Protein folding under confinement: A role for solvent

    PubMed Central

    Lucent, Del; Vishal, V.; Pande, Vijay S.

    2007-01-01

    Although most experimental and theoretical studies of protein folding involve proteins in vitro, the effects of spatial confinement may complicate protein folding in vivo. In this study, we examine the folding dynamics of villin (a small fast folding protein) with explicit solvent confined to an inert nanopore. We have calculated the probability of folding before unfolding (Pfold) under various confinement regimes. Using Pfold correlation techniques, we observed two competing effects. Confining protein alone promotes folding by destabilizing the unfolded state. In contrast, confining both protein and solvent gives rise to a solvent-mediated effect that destabilizes the native state. When both protein and solvent are confined we see unfolding to a compact unfolded state different from the unfolded state seen in bulk. Thus, we demonstrate that the confinement of solvent has a significant impact on protein kinetics and thermodynamics. We conclude with a discussion of the implications of these results for folding in confined environments such as the chaperonin cavity in vivo. PMID:17563390

  16. A hybrid MD-kMC algorithm for folding proteins in explicit solvent.

    PubMed

    Peter, Emanuel Karl; Shea, Joan-Emma

    2014-04-14

    We present a novel hybrid MD-kMC algorithm that is capable of efficiently folding proteins in explicit solvent. We apply this algorithm to the folding of a small protein, Trp-Cage. Different kMC move sets that capture different possible rate limiting steps are implemented. The first uses secondary structure formation as a relevant rate event (a combination of dihedral rotations and hydrogen-bonding formation and breakage). The second uses tertiary structure formation events through formation of contacts via translational moves. Both methods fold the protein, but via different mechanisms and with different folding kinetics. The first method leads to folding via a structured helical state, with kinetics fit by a single exponential. The second method leads to folding via a collapsed loop, with kinetics poorly fit by single or double exponentials. In both cases, folding times are faster than experimentally reported values, The secondary and tertiary move sets are integrated in a third MD-kMC implementation, which now leads to folding of the protein via both pathways, with single and double-exponential fits to the rates, and to folding rates in good agreement with experimental values. The competition between secondary and tertiary structure leads to a longer search for the helix-rich intermediate in the case of the first pathway, and to the emergence of a kinetically trapped long-lived molten-globule collapsed state in the case of the second pathway. The algorithm presented not only captures experimentally observed folding intermediates and kinetics, but yields insights into the relative roles of local and global interactions in determining folding mechanisms and rates. PMID:24499973

  17. A hierarchical protein folding scheme based on the building block folding model.

    PubMed

    Haspel, Nurit; Wainreb, Gilad; Inbar, Yuval; Tsai, Hui-Hsu; Tsai, Chung-Jung; Wolfson, Haim J; Nussinov, Ruth

    2007-01-01

    The building block protein folding model states that the native protein structure is the product of a combinatorial assembly of relatively structurally independent contiguous parts of the protein that possess a hydrophobic core, i.e., building blocks (BBs). According to this model, our group proposed a three-stage scheme for a feasible time-wise semi ab-intio protein structure prediction. Given a protein sequence, at the first stage of the prediction scheme, we propose cutting the sequence into structurally assigned BBs. Next, we perform a combinatorial assembly and attempt to predict the relative three-dimensional arrangement of the BBs. In the third stage, we refine and rank the assemblies. The scheme has proven to be very promising in reducing the complexity of the protein folding problem and gaining insight into the protein folding process. In this chapter, we describe the different stages of the scheme and discuss a possible application of the model to protein design. PMID:16957324

  18. Kinetic Atom.

    ERIC Educational Resources Information Center

    Wilson, David B.

    1981-01-01

    Surveys the research of scientists like Joule, Kelvin, Maxwell, Clausius, and Boltzmann as it comments on the basic conceptual issues involved in the development of a more precise kinetic theory and the idea of a kinetic atom. (Author/SK)

  19. Enzyme Kinetics.

    ERIC Educational Resources Information Center

    Moe, Owen; Cornelius, Richard

    1988-01-01

    Conveys an appreciation of enzyme kinetic analysis by using a practical and intuitive approach. Discusses enzyme assays, kinetic models and rate laws, the kinetic constants (V, velocity, and Km, Michaels constant), evaluation of V and Km from experimental data, and enzyme inhibition. (CW)

  20. Rapid compaction during RNA folding

    NASA Astrophysics Data System (ADS)

    Russell, Rick; Millett, Ian S.; Tate, Mark W.; Kwok, Lisa W.; Nakatani, Bradley; Gruner, Sol M.; Mochrie, Simon G. J.; Pande, Vijay; Doniach, Sebastian; Herschlag, Daniel; Pollack, Lois

    2002-04-01

    We have used small angle x-ray scattering and computer simulations with a coarse-grained model to provide a time-resolved picture of the global folding process of the Tetrahymena group I RNA over a time window of more than five orders of magnitude. A substantial phase of compaction is observed on the low millisecond timescale, and the overall compaction and global shape changes are largely complete within one second, earlier than any known tertiary contacts are formed. This finding indicates that the RNA forms a nonspecifically collapsed intermediate and then searches for its tertiary contacts within a highly restricted subset of conformational space. The collapsed intermediate early in folding of this RNA is grossly akin to molten globule intermediates in protein folding.

  1. Globular Protein Folding In Vitro and In Vivo.

    PubMed

    Gruebele, Martin; Dave, Kapil; Sukenik, Shahar

    2016-07-01

    In vitro, computational, and theoretical studies of protein folding have converged to paint a rich and complex energy landscape. This landscape is sensitively modulated by environmental conditions and subject to evolutionary pressure on protein function. Of these environments, none is more complex than the cell itself, where proteins function in the cytosol, in membranes, and in different compartments. A wide variety of kinetic and thermodynamics experiments, ranging from single-molecule studies to jump kinetics and from nuclear magnetic resonance to imaging on the microscope, have elucidated how protein energy landscapes facilitate folding and how they are subject to evolutionary constraints and environmental perturbation. Here we review some recent developments in the field and refer the reader to some original work and additional reviews that cover this broad topic in protein science. PMID:27391927

  2. NoFold: RNA structure clustering without folding or alignment.

    PubMed

    Middleton, Sarah A; Kim, Junhyong

    2014-11-01

    Structures that recur across multiple different transcripts, called structure motifs, often perform a similar function-for example, recruiting a specific RNA-binding protein that then regulates translation, splicing, or subcellular localization. Identifying common motifs between coregulated transcripts may therefore yield significant insight into their binding partners and mechanism of regulation. However, as most methods for clustering structures are based on folding individual sequences or doing many pairwise alignments, this results in a tradeoff between speed and accuracy that can be problematic for large-scale data sets. Here we describe a novel method for comparing and characterizing RNA secondary structures that does not require folding or pairwise alignment of the input sequences. Our method uses the idea of constructing a distance function between two objects by their respective distances to a collection of empirical examples or models, which in our case consists of 1973 Rfam family covariance models. Using this as a basis for measuring structural similarity, we developed a clustering pipeline called NoFold to automatically identify and annotate structure motifs within large sequence data sets. We demonstrate that NoFold can simultaneously identify multiple structure motifs with an average sensitivity of 0.80 and precision of 0.98 and generally exceeds the performance of existing methods. We also perform a cross-validation analysis of the entire set of Rfam families, achieving an average sensitivity of 0.57. We apply NoFold to identify motifs enriched in dendritically localized transcripts and report 213 enriched motifs, including both known and novel structures.

  3. Folding analysis of the most complex Stevedore's protein knot.

    PubMed

    Wang, Iren; Chen, Szu-Yu; Hsu, Shang-Te Danny

    2016-01-01

    DehI is a homodimeric haloacid dehalogenase from Pseudomonas putida that contains the most complex 61 Stevedore's protein knot within its folding topology. To examine how DehI attains such an intricate knotted topology we combined far-UV circular dichroism (CD), intrinsic fluorescence spectroscopy and small angle X-ray scattering (SAXS) to investigate its folding mechanism. Equilibrium unfolding of DehI by chemical denaturation indicated the presence of two highly populated folding intermediates, I and I'. While the two intermediates vary in secondary structure contents and tertiary packing according to CD and intrinsic fluorescence, respectively, their overall dimension and compactness are similar according to SAXS. Three single-tryptophan variants (W34, W53, and W196) were generated to probe non-cooperative unfolding events localized around the three fluorophores. Kinetic fluorescence measurements indicated that the transition from the intermediate I' to the unfolded state is rate limiting. Our multiparametric folding analyses suggest that DehI unfolds through a linear folding pathway with two distinct folding intermediates by initial hydrophobic collapse followed by nucleation condensation, and that knotting precedes the formation of secondary structures. PMID:27527519

  4. Semiempirical prediction of protein folds

    NASA Astrophysics Data System (ADS)

    Fernández, Ariel; Colubri, Andrés; Appignanesi, Gustavo

    2001-08-01

    We introduce a semiempirical approach to predict ab initio expeditious pathways and native backbone geometries of proteins that fold under in vitro renaturation conditions. The algorithm is engineered to incorporate a discrete codification of local steric hindrances that constrain the movements of the peptide backbone throughout the folding process. Thus, the torsional state of the chain is assumed to be conditioned by the fact that hopping from one basin of attraction to another in the Ramachandran map (local potential energy surface) of each residue is energetically more costly than the search for a specific (Φ, Ψ) torsional state within a single basin. A combinatorial procedure is introduced to evaluate coarsely defined torsional states of the chain defined ``modulo basins'' and translate them into meaningful patterns of long range interactions. Thus, an algorithm for structure prediction is designed based on the fact that local contributions to the potential energy may be subsumed into time-evolving conformational constraints defining sets of restricted backbone geometries whereupon the patterns of nonbonded interactions are constructed. The predictive power of the algorithm is assessed by (a) computing ab initio folding pathways for mammalian ubiquitin that ultimately yield a stable structural pattern reproducing all of its native features, (b) determining the nucleating event that triggers the hydrophobic collapse of the chain, and (c) comparing coarse predictions of the stable folds of moderately large proteins (N~100) with structural information extracted from the protein data bank.

  5. Direct molecular dynamics observation of protein folding transition state ensemble.

    PubMed Central

    Ding, Feng; Dokholyan, Nikolay V; Buldyrev, Sergey V; Stanley, H Eugene; Shakhnovich, Eugene I

    2002-01-01

    The concept of the protein transition state ensemble (TSE), a collection of the conformations that have 50% probability to convert rapidly to the folded state and 50% chance to rapidly unfold, constitutes the basis of the modern interpretation of protein engineering experiments. It has been conjectured that conformations constituting the TSE in many proteins are the expanded and distorted forms of the native state built around a specific folding nucleus. This view has been supported by a number of on-lattice and off-lattice simulations. Here we report a direct observation and characterization of the TSE by molecular dynamic folding simulations of the C-Src SH3 domain, a small protein that has been extensively studied experimentally. Our analysis reveals a set of key interactions between residues, conserved by evolution, that must be formed to enter the kinetic basin of attraction of the native state. PMID:12496119

  6. Microwave-enhanced folding and denaturation of globular proteins

    NASA Astrophysics Data System (ADS)

    Bohr, Henrik; Bohr, Jakob

    2000-04-01

    It is shown that microwave irradiation can affect the kinetics of the folding process of some globular proteins, especially β-lactoglobulin. At low temperature the folding from the cold denatured phase of the protein is enhanced, while at a higher temperature the denaturation of the protein from its folded state is enhanced. In the latter case, a negative temperature gradient is needed for the denaturation process, suggesting that the effects of the microwaves are nonthermal. This supports the notion that coherent topological excitations can exist in proteins. The application of microwaves hold promises for a wide range of biotechnological applications, such as protein synthesis, protein aggregation, etc., and may have implications for biological systems as well.

  7. Slowing Down Downhill Folding: A Three-Probe Study

    PubMed Central

    Kim, Seung Joong; Matsumura, Yoshitaka; Dumont, Charles; Kihara, Hiroshi; Gruebele, Martin

    2009-01-01

    Abstract The mutant Tyr22Trp/Glu33Tyr/Gly46Ala/Gly48Ala of λ repressor fragment λ6−85 was previously assigned as an incipient downhill folder. We slow down its folding in a cryogenic water-ethylene-glycol solvent (−18 to −28°C). The refolding kinetics are probed by small-angle x-ray scattering, circular dichroism, and fluorescence to measure the radius of gyration, the average secondary structure content, and the native packing around the single tryptophan residue. The main resolved kinetic phase of the mutant is probe independent and faster than the main phase observed for the pseudo-wild-type. Excess helical structure formed early on by the mutant may reduce the formation of turns and prevent the formation of compact misfolded states, speeding up the overall folding process. Extrapolation of our main cryogenic folding phase and previous T-jump measurements to 37°C yields nearly the same refolding rate as extrapolated by Oas and co-workers from NMR line-shape data. Taken together, all the data consistently indicate a folding speed limit of ∼4.5 μs for this fast folder. PMID:19580767

  8. A simple quantitative model of macromolecular crowding effects on protein folding: Application to the murine prion protein(121-231)

    NASA Astrophysics Data System (ADS)

    Bergasa-Caceres, Fernando; Rabitz, Herschel A.

    2013-06-01

    A model of protein folding kinetics is applied to study the effects of macromolecular crowding on protein folding rate and stability. Macromolecular crowding is found to promote a decrease of the entropic cost of folding of proteins that produces an increase of both the stability and the folding rate. The acceleration of the folding rate due to macromolecular crowding is shown to be a topology-dependent effect. The model is applied to the folding dynamics of the murine prion protein (121-231). The differential effect of macromolecular crowding as a function of protein topology suffices to make non-native configurations relatively more accessible.

  9. Chemical methods for producing disulfide bonds in peptides and proteins to study folding regulation.

    PubMed

    Okumura, Masaki; Shimamoto, Shigeru; Hidaka, Yuji

    2014-04-01

    Disulfide bonds play a critical role in the folding of secretory and membrane proteins. Oxidative folding reactions of disulfide bond-containing proteins typically require several hours or days, and numerous misbridged disulfide isomers are often observed as intermediates. The rate-determining step in refolding is thought to be the disulfide-exchange reaction from nonnative to native disulfide bonds in folding intermediates, which often precipitate during the refolding process because of their hydrophobic properties. To overcome this, chemical additives or a disulfide catalyst, protein disulfide isomerase (PDI), are generally used in refolding experiments to regulate disulfide-coupled peptide and protein folding. This unit describes such methods in the context of the thermodynamic and kinetic control of peptide and protein folding, including (1) regulation of disulfide-coupled peptides and protein folding assisted by chemical additives, (2) reductive unfolding of disulfide-containing peptides and proteins, and (3) regulation of disulfide-coupled peptide and protein folding using PDI.

  10. Substrate-induced activation of a trapped IMC-mediated protein folding intermediate.

    PubMed

    Inouye, M; Fu, X; Shinde, U

    2001-04-01

    While several unfolded proteins acquire native structures through distinct folding intermediates, the physiological relevance and importance of such states in the folding kinetics remain controversial. The intramolecular chaperone (IMC) of subtilisin was used to trap a partially folded, stable crosslinked intermediate conformer (CLIC) through a disulfide bond between mutated IMC and subtilisin. The trapped CLIC contains non-native interactions. Here we show that CLIC can be induced into a catalytically active form by incubating it with small peptide substrates. The structure and catalytic properties of the activated crosslinked intermediate conformer (A-CLIC) differ from those of the fully folded enzyme in that A-CLIC lacks any endopeptidase activity toward a large protein substrate. Our results show that a disulfide-linked partially folded protein can be induced to acquire catalytic activity with a substrate specificity that is different from completely folded subtilisin. These results also suggest that protein folding intermediates may also participate in catalytic reactions.

  11. Folding of a large protein at high structural resolution

    PubMed Central

    Walters, Benjamin T.; Mayne, Leland; Hinshaw, James R.; Sosnick, Tobin R.; Englander, S. Walter

    2013-01-01

    Kinetic folding of the large two-domain maltose binding protein (MBP; 370 residues) was studied at high structural resolution by an advanced hydrogen-exchange pulse-labeling mass-spectrometry method (HX MS). Dilution into folding conditions initiates a fast molecular collapse into a polyglobular conformation (<20 ms), determined by various methods including small angle X-ray scattering. The compaction produces a structurally heterogeneous state with widespread low-level HX protection and spectroscopic signals that match the equilibrium melting posttransition-state baseline. In a much slower step (7-s time constant), all of the MBP molecules, although initially heterogeneously structured, form the same distinct helix plus sheet folding intermediate with the same time constant. The intermediate is composed of segments that are distant in the MBP sequence but adjacent in the native protein where they close the longest residue-to-residue contact. Segments that are most HX protected in the early molecular collapse do not contribute to the initial intermediate, whereas the segments that do participate are among the less protected. The 7-s intermediate persists through the rest of the folding process. It contains the sites of three previously reported destabilizing mutations that greatly slow folding. These results indicate that the intermediate is an obligatory step on the MBP folding pathway. MBP then folds to the native state on a longer time scale (∼100 s), suggestively in more than one step, the first of which forms structure adjacent to the 7-s intermediate. These results add a large protein to the list of proteins known to fold through distinct native-like intermediates in distinct pathways. PMID:24191053

  12. Statistical Mechanical Model for pH-Induced Protein Folding: Application to Apomyoglobin.

    PubMed

    Mizukami, Takuya; Sakuma, Yosuke; Maki, Kosuke

    2016-09-01

    Despite the major role of pH in protein folding and stability, a quantitative understanding of the pH-induced protein folding mechanism remains elusive. Two conventional models, the Monod-Wyman-Changeux and Linderstrøm-Lang smeared charge models, respectively, have been used to analyze the formation/disruption of specific native structures and fluctuating non-native states. However, there are only a few models that can represent the overall kinetic events of folding/unfolding independent of the properties of relevant molecular species, which has hampered the efforts to systematically analyze pH-induced folding. Here, we constructed a statistical mechanical model that incorporates the protonation mechanism of conventional models along with a combined manual search and least-squares fitting procedure, which was used to investigate the folding of horse apomyoglobin over a wide pH range (2.2-6.7), with a time window ranging from ∼40 μs to ∼100 s, using continuous-/stopped-flow fluorescence at 8 °C. Quantitative analysis assuming a five-state sequential scheme indicated that (1) pH-induced folding/unfolding is represented by both specific binding and Coulombic interactions; (2) kinetic folding/unfolding intermediates share kinetic mechanisms with the equilibrium intermediate, indicating their equivalence; and (3) native-like properties are acquired successively during folding by intermediates and in transition states. This model could also be applied to a variety of association/dissociation processes. PMID:27491483

  13. A magnetization-transfer nuclear magnetic resonance study of the folding of staphylococcal nuclease

    SciTech Connect

    Evans, P.A.; Kautz, R.A.; Fox, R.O.; Dobson, C.M. )

    1989-01-10

    The equilibrium between alternative folded states of a globular protein, staphylococcal nuclease, has been investigated by using {sup 1}H NMR. Magnetization-transfer experiments have revealed the existence of a related structural heterogeneity of the unfolded state, and quantitative analysis of a series of these experiments has permitted the kinetics of folding and interconversion of the different states to be explored. A model based on cis/trans isomerism at the peptide bond preceding Pro-117 has been developed to account for the results. This model, recently supported by a protein-engineering experiment has been used to interpret the kinetic data, providing insight into the nature of the folding processes. The predominance of the cis-proline form in the folded state is shown to derive from a large favorable enthalpy term resulting from more effective overall folding interactions. The kinetics of folding and isomerization are shown to occur on similar time scales, such that more than one pathway between two states may be significant. It has been possible, however, to compare the direct folding and unfolding rates within the cis- and trans-proline-containing populations, with results suggesting that the specific stabilization of the cis peptide bond is effective only at a late stage in the folding process.

  14. RNAiFold: a web server for RNA inverse folding and molecular design

    PubMed Central

    Garcia-Martin, Juan Antonio; Clote, Peter; Dotu, Ivan

    2013-01-01

    Synthetic biology and nanotechnology are poised to make revolutionary contributions to the 21st century. In this article, we describe a new web server to support in silico RNA molecular design. Given an input target RNA secondary structure, together with optional constraints, such as requiring GC-content to lie within a certain range, requiring the number of strong (GC), weak (AU) and wobble (GU) base pairs to lie in a certain range, the RNAiFold web server determines one or more RNA sequences, whose minimum free-energy secondary structure is the target structure. RNAiFold provides access to two servers: RNA-CPdesign, which applies constraint programming, and RNA-LNSdesign, which applies the large neighborhood search heuristic; hence, it is suitable for larger input structures. Both servers can also solve the RNA inverse hybridization problem, i.e. given a representation of the desired hybridization structure, RNAiFold returns two sequences, whose minimum free-energy hybridization is the input target structure. The web server is publicly accessible at http://bioinformatics.bc.edu/clotelab/RNAiFold, which provides access to two specialized servers: RNA-CPdesign and RNA-LNSdesign. Source code for the underlying algorithms, implemented in COMET and supported on linux, can be downloaded at the server website. PMID:23700314

  15. RNAiFold: a web server for RNA inverse folding and molecular design.

    PubMed

    Garcia-Martin, Juan Antonio; Clote, Peter; Dotu, Ivan

    2013-07-01

    Synthetic biology and nanotechnology are poised to make revolutionary contributions to the 21st century. In this article, we describe a new web server to support in silico RNA molecular design. Given an input target RNA secondary structure, together with optional constraints, such as requiring GC-content to lie within a certain range, requiring the number of strong (GC), weak (AU) and wobble (GU) base pairs to lie in a certain range, the RNAiFold web server determines one or more RNA sequences, whose minimum free-energy secondary structure is the target structure. RNAiFold provides access to two servers: RNA-CPdesign, which applies constraint programming, and RNA-LNSdesign, which applies the large neighborhood search heuristic; hence, it is suitable for larger input structures. Both servers can also solve the RNA inverse hybridization problem, i.e. given a representation of the desired hybridization structure, RNAiFold returns two sequences, whose minimum free-energy hybridization is the input target structure. The web server is publicly accessible at http://bioinformatics.bc.edu/clotelab/RNAiFold, which provides access to two specialized servers: RNA-CPdesign and RNA-LNSdesign. Source code for the underlying algorithms, implemented in COMET and supported on linux, can be downloaded at the server website.

  16. Folded supersymmetry with a twist

    DOE PAGESBeta

    Cohen, Timothy; Craig, Nathaniel; Lou, Hou Keong; Pinner, David

    2016-03-30

    Folded supersymmetry (f-SUSY) stabilizes the weak scale against radiative corrections from the top sector via scalar partners whose gauge quantum numbers differ from their Standard Model counterparts. This non-trivial pairing of states can be realized in extra-dimensional theories with appropriate supersymmetry-breaking boundary conditions. We present a class of calculable f-SUSY models that are parametrized by a non-trivial twist in 5D boundary conditions and can accommodate the observed Higgs mass and couplings. Although the distinctive phenomenology associated with the novel folded states should provide strong evidence for this mechanism, the most stringent constraints are currently placed by conventional supersymmetry searches. Asmore » a result, these models remain minimally fine-tuned in light of LHC8 data and provide a range of both standard and exotic signatures accessible at LHC13.« less

  17. Mechanical Folding and Unfolding of Protein Barnase at the Single-Molecule Level.

    PubMed

    Alemany, Anna; Rey-Serra, Blanca; Frutos, Silvia; Cecconi, Ciro; Ritort, Felix

    2016-01-01

    The unfolding and folding of protein barnase has been extensively investigated in bulk conditions under the effect of denaturant and temperature. These experiments provided information about structural and kinetic features of both the native and the unfolded states of the protein, and debates about the possible existence of an intermediate state in the folding pathway have arisen. Here, we investigate the folding/unfolding reaction of protein barnase under the action of mechanical force at the single-molecule level using optical tweezers. We measure unfolding and folding force-dependent kinetic rates from pulling and passive experiments, respectively, and using Kramers-based theories (e.g., Bell-Evans and Dudko-Hummer-Szabo models), we extract the position of the transition state and the height of the kinetic barrier mediating unfolding and folding transitions, finding good agreement with previous bulk measurements. Measurements of the force-dependent kinetic barrier using the continuous effective barrier analysis show that protein barnase verifies the Leffler-Hammond postulate under applied force and allow us to extract its free energy of folding, ΔG0. The estimated value of ΔG0 is in agreement with our predictions obtained using fluctuation relations and previous bulk studies. To address the possible existence of an intermediate state on the folding pathway, we measure the power spectrum of force fluctuations at high temporal resolution (50 kHz) when the protein is either folded or unfolded and, additionally, we study the folding transition-path time at different forces. The finite bandwidth of our experimental setup sets the lifetime of potential intermediate states upon barnase folding/unfolding in the submillisecond timescale. PMID:26745410

  18. Application of Time-Resolved Tryptophan Phosphorescence Spectroscopy to Protein Folding Studies.

    NASA Astrophysics Data System (ADS)

    Subramaniam, Vinod

    This thesis presents studies of the protein folding problem, one of the most significant questions in contemporary biophysics. Sensitive biophysical techniques, including room temperature tryptophan phosphorescence, which reports on the local environment of the residue, and the lability of proteins to denaturation, a global parameter, were used to assess the validity of the traditional assumption that the biologically active state of a protein is the 'native' state, and to determine whether the pathways of folding in vitro lead to the folded state achieved in vivo. Phosphorescence techniques have also been extended to study, for the first time, emission from tryptophan residues engineered into specific positions as reporters of protein structure. During in vitro refolding of E. coli alkaline phosphatase and bovine 13-lactoglobulin, significant differences were found between the refolded proteins and the native conformations, which have no apparent effect on the biological functions. Slow conformational transitions, termed 'annealing,' that occur long after the return of enzyme activity of alkaline phosphatase are manifested in the retarded recovery of phosphorescence intensity, lifetime, and protein lability. While 'annealing' is not observed for beta -lactoglobulin, both phosphorescence and lability experiments reveal changes in the structure of the refolded protein, even though its biological activity, retinol binding, is fully recovered. This result suggests that the pathways of folding in vitro need not lead to the structure formed in vivo. We have used phosphorescence techniques to study the refolding of ribonuclease T1, which exhibits slow kinetics characteristic of proline isomerization. Furthermore, the ability to extract structural information from phosphorescent tryptophan probes engineered into selected regions represents an important advance in studying protein structure; we have reported the first such results from a mutant staphylococcal nuclease. The

  19. Kinetic: A system code for analyzing nuclear thermal propulsion rocket engine transients

    NASA Technical Reports Server (NTRS)

    Schmidt, Eldon; Lazareth, Otto; Ludewig, Hans

    1993-01-01

    The topics are presented in viewgraph form and include the following: outline of kinetic code; a kinetic information flow diagram; kinetic neutronic equations; turbopump/nozzle algorithm; kinetic heat transfer equations per node; and test problem diagram.

  20. Chain length scaling of protein folding time: Beta sheet structures

    NASA Astrophysics Data System (ADS)

    Dimitrievski, K.; Kasemo, B.; Zhdanov, V. P.

    2000-07-01

    We present comprehensive 3D lattice Monte Carlo simulations of the folding kinetics of two-turn antiparallel β sheets. The model employed takes into account isotropic nonspecific interactions as in previous flexible heteropolymer models and also orientation-dependent monomer-monomer interactions, mimicking the formation of hydrogen bonds and chain rigidity. The chain length is varied from N=15 to 33. For each chain length, we calculate the fastest folding temperature, Tfast, folding temperature, Tfold, and glass-transition temperature, Tg. The time-averaged occupation probability of the native state is found to be nearly independent of N at all temperatures. The dependence of Tfast and Tfold on N is accordingly relatively weak. The temperature interval where the folding is fast rapidly decreases with increasing N. For the chain lengths chosen, Tfold slightly exceeds Tg. The dependence of the folding time τf on N is well fitted by using the power law, τf∝Nλ. The exponent λ is found to depend on temperature and on the distribution of nonspecific interactions in the chain. In particular, λ=2.7-4.0 at T=Tfast and 5.2 at T slightly below Tfold. Evaluating τf in real units at T near Tfold yields physically reasonable results.

  1. Complete RNA inverse folding: computational design of functional hammerhead ribozymes

    PubMed Central

    Dotu, Ivan; Garcia-Martin, Juan Antonio; Slinger, Betty L.; Mechery, Vinodh; Meyer, Michelle M.; Clote, Peter

    2014-01-01

    Nanotechnology and synthetic biology currently constitute one of the most innovative, interdisciplinary fields of research, poised to radically transform society in the 21st century. This paper concerns the synthetic design of ribonucleic acid molecules, using our recent algorithm, RNAiFold, which can determine all RNA sequences whose minimum free energy secondary structure is a user-specified target structure. Using RNAiFold, we design ten cis-cleaving hammerhead ribozymes, all of which are shown to be functional by a cleavage assay. We additionally use RNAiFold to design a functional cis-cleaving hammerhead as a modular unit of a synthetic larger RNA. Analysis of kinetics on this small set of hammerheads suggests that cleavage rate of computationally designed ribozymes may be correlated with positional entropy, ensemble defect, structural flexibility/rigidity and related measures. Artificial ribozymes have been designed in the past either manually or by SELEX (Systematic Evolution of Ligands by Exponential Enrichment); however, this appears to be the first purely computational design and experimental validation of novel functional ribozymes. RNAiFold is available at http://bioinformatics.bc.edu/clotelab/RNAiFold/. PMID:25209235

  2. Intermediates and the folding of proteins L and G

    SciTech Connect

    Brown, Scott; Head-Gordon, Teresa

    2003-07-01

    We use a minimalist protein model, in combination with a sequence design strategy, to determine differences in primary structure for proteins L and G that are responsible for the two proteins folding through distinctly different folding mechanisms. We find that the folding of proteins L and G are consistent with a nucleation-condensation mechanism, each of which is described as helix-assisted {beta}-1 and {beta}-2 hairpin formation, respectively. We determine that the model for protein G exhibits an early intermediate that precedes the rate-limiting barrier of folding and which draws together misaligned secondary structure elements that are stabilized by hydrophobic core contacts involving the third {beta}-strand, and presages the later transition state in which the correct strand alignment of these same secondary structure elements is restored. Finally the validity of the targeted intermediate ensemble for protein G was analyzed by fitting the kinetic data to a two-step first order reversible reaction, proving that protein G folding involves an on-pathway early intermediate, and should be populated and therefore observable by experiment.

  3. Ventricular-Fold Dynamics in Human Phonation

    ERIC Educational Resources Information Center

    Bailly, Lucie; Bernardoni, Nathalie Henrich; Müller, Frank; Rohlfs, Anna-Katharina; Hess, Markus

    2014-01-01

    Purpose: In this study, the authors aimed (a) to provide a classification of the ventricular-fold dynamics during voicing, (b) to study the aerodynamic impact of these motions on vocal-fold vibrations, and (c) to assess whether ventricular-fold oscillations could be sustained by aerodynamic coupling with the vocal folds. Method: A 72-sample…

  4. The radius of gyration of an apomyoglobin folding intermediate

    SciTech Connect

    Eliezer, D.; Jennings, P.A.; Wright, P.E.

    1995-10-20

    Apomyoglobin (apoMb) forms a stable compact partially folded state under acidic conditions. This {open_quotes}molten globule{close_quotes} intermediate is slightly expanded relative to the native form of the protein, with a radius of gyration (R{sub g}) of 23 ({plus_minus} 2) {Angstrom} versus 19 ({plus_minus}) {Angstrom}, and shows stable secondary structure in the A,G, and H helices. We demonstrated recently, with the use of stopped-flow circular dichroism and pulse-labeling hydrogen exchange measurements, that the earliest detectable intermediate (formed with 6 ms) in the apoMb kinetic refolding pathway closely resembles the equilibrium molten globule state populated under acid conditions. A key question remained as to how compact this kinetic intermediate is compared to the equilibrium and native states. The cooperative unfolding of the kinetic intermediate and the significant protection from amide proton exchange (as compared to corresponding isolated peptides in solution) led us to propose that the kinetic intermediate is also compact. Such a proposal could best be verified by direct determination of the size of the protein as it folds, but measurements of this nature were not feasible at the time. Newly developed improvements in time-resolved small angle x-ray scattering (SAXS) experiments allow direct measurement of the time-dependent change of R{sub g} of a protein as it folds in the millisecond to second time frame. We initiated studies of the refolding of apoMb using this technique, under conditions similar to those employed in our previous work. SAXS data collected during the first 100 ms after initiation of the refolding reaction are shown. 11 refs., 2 figs.

  5. Inflation Rates, Car Devaluation, and Chemical Kinetics.

    ERIC Educational Resources Information Center

    Pogliani, Lionello; Berberan-Santos, Mario N.

    1996-01-01

    Describes the inflation rate problem and offers an interesting analogy with chemical kinetics. Presents and solves the car devaluation problem as a normal chemical kinetic problem where the order of the rate law and the value of the rate constant are derived. (JRH)

  6. Exploring the folding pathway of green fluorescent protein through disulfide engineering.

    PubMed

    Pitman, Derek J; Banerjee, Shounak; Macari, Stephen J; Castaldi, Christopher A; Crone, Donna E; Bystroff, Christopher

    2015-03-01

    We have introduced two disulfide crosslinks into the loop regions on opposite ends of the beta barrel in superfolder green fluorescent protein (GFP) in order to better understand the nature of its folding pathway. When the disulfide on the side opposite the N/C-termini is formed, folding is 2× faster, unfolding is 2000× slower, and the protein is stabilized by 16 kJ/mol. But when the disulfide bond on the side of the termini is formed we see little change in the kinetics and stability. The stabilization upon combining the two crosslinks is approximately additive. When the kinetic effects are broken down into multiple phases, we observe Hammond behavior in the upward shift of the kinetic m-value of unfolding. We use these results in conjunction with structural analysis to assign folding intermediates to two parallel folding pathways. The data are consistent with a view that the two fastest transition states of folding are "barrel closing" steps. The slower of the two phases passes through an intermediate with the barrel opening occurring between strands 7 and 8, while the faster phase opens between 9 and 4. We conclude that disulfide crosslink-induced perturbations in kinetics are useful for mapping the protein folding pathway.

  7. Folding of a Cyclin Box

    PubMed Central

    Chemes, Lucía B.; Noval, María G.; Sánchez, Ignacio E.; de Prat-Gay, Gonzalo

    2013-01-01

    The retinoblastoma tumor suppressor (Rb) controls the proliferation, differentiation, and survival of cells in most eukaryotes with a role in the fate of stem cells. Its inactivation by mutation or oncogenic viruses is required for cellular transformation and eventually carcinogenesis. The high conservation of the Rb cyclin fold prompted us to investigate the link between conformational stability and ligand binding properties of the RbAB pocket domain. RbAB unfolding presents a three-state transition involving cooperative secondary and tertiary structure changes and a partially folded intermediate that can oligomerize. The first transition corresponds to unfolding of the metastable B subdomain containing the binding site for the LXCXE motif present in cellular and viral targets, and the second transition corresponds to the stable A subdomain. The low thermodynamic stability of RbAB translates into a propensity to rapidly oligomerize and aggregate at 37 °C (T50 = 28 min) that is suppressed by human papillomavirus E7 and E2F peptide ligands, suggesting that Rb is likely stabilized in vivo through binding to target proteins. We propose that marginal stability and associated oligomerization may be conserved for function as a “hub” protein, allowing the formation of multiprotein complexes, which could constitute a robust mechanism to retain its cell cycle regulatory role throughout evolution. Decreased stability and oligomerization are shared with the p53 tumor suppressor, suggesting a link between folding and function in these two essential cell regulators that are inactivated in most cancers and operate within multitarget signaling pathways. PMID:23632018

  8. Landscapes with megabasins: Polyamorphism in liquids and biopolymers and the role of nucleation in folding and folding diseases

    NASA Astrophysics Data System (ADS)

    Angell, C. A.

    1997-02-01

    We show how energy landscape concepts can rationalize the observations on glassforming liquids over the whole range of behavior, strong to fragile. In particular, we show how the existence of landscapes with both strong and fragile megabasins can provide a basis for understanding the nature of quasi-first-order transitions between amorphous states such as those observed to occur in the glassy states of “strong” glassformers. We show how this propensity originates in the liquid state and then emphasize the analogy provided, at the mesoscopic level, by the folding transition in proteins. Recognition that the folding transition is an equilibrium first-order transition between polyamorphic forms of a complex system implies recognition of the need for a nucleation step in the process. When nucleated phase transitions are kinetically retarded, their probability can be influenced by time-temperature history and by the presence of nucleating agents. Nucleation events are statistically rare in mesoscopic systems, hence the ability to fold rapidly should require special features in the folding molecular structure or the presence of nucleating agents. We propose that the unwanted folding events leading to pathogenic forms of certain proteins (prions) can be stimulated by nucleating agents, which thus may be the unidentified infectious agents in “mad cow” disease and related maladies.

  9. Chaperonin-mediated Protein Folding

    PubMed Central

    Horwich, Arthur L.

    2013-01-01

    We have been studying chaperonins these past twenty years through an initial discovery of an action in protein folding, analysis of structure, and elucidation of mechanism. Some of the highlights of these studies were presented recently upon sharing the honor of the 2013 Herbert Tabor Award with my early collaborator, Ulrich Hartl, at the annual meeting of the American Society for Biochemistry and Molecular Biology in Boston. Here, some of the major findings are recounted, particularly recognizing my collaborators, describing how I met them and how our great times together propelled our thinking and experiments. PMID:23803606

  10. Folding of horse cytochrome c in the reduced state.

    PubMed

    Bhuyan, A K; Udgaonkar, J B

    2001-10-01

    Equilibrium and kinetic folding studies of horse cytochrome c in the reduced state have been carried out under strictly anaerobic conditions at neutral pH, 10 degrees C, in the entire range of aqueous solubility of guanidinium hydrochloride (GdnHCl). Equilibrium unfolding transitions observed by Soret heme absorbance, excitation energy transfer from the lone tryptophan residue to the ferrous heme, and far-UV circular dichroism (CD) are all biphasic and superimposable, implying no accumulation of structural intermediates. The thermodynamic parameters obtained by two-state analysis of these transitions yielded DeltaG(H2O)=18.8(+/-1.45) kcal mol(-1), and C(m)=5.1(+/-0.15) M GdnHCl, indicating unusual stability of reduced cytochrome c. These results have been used in conjunction with the redox potential of native cytochrome c and the known stability of oxidized cytochrome c to estimate a value of -164 mV as the redox potential of the unfolded protein. Stopped-flow kinetics of folding and unfolding have been recorded by Soret heme absorbance, and tryptophan fluorescence as observables. The refolding kinetics are monophasic in the transition region, but become biphasic as moderate to strongly native-like conditions are approached. There also is a burst folding reaction unobservable in the stopped-flow time window. Analyses of the two observable rates and their amplitudes indicate that the faster of the two rates corresponds to apparent two-state folding (U<-->N) of 80-90 % of unfolded molecules with a time constant in the range 190-550 micros estimated by linear extrapolation and model calculations. The remaining 10-20 % of the population folds to an off-pathway intermediate, I, which is required to unfold first to the initial unfolded state, U, in order to refold correctly to the native state, N (I<-->U<-->N). The slower of the two observable rates, which has a positive slope in the linear functional dependence on the denaturant concentration indicating that an

  11. Fold-A-Board communication device.

    PubMed

    Sokolowski, Joanna; Maher, Aedan; Jaycox, Katie; Siegler, Kevin

    2011-01-01

    Aphasia is an impairment of language resulting from stroke that can affect a person's ability to use and comprehend words. People who have aphasia often keep a writing device handy to write down information that they have trouble communicating verbally. Device options include expensive keyboard devices or more commonly small inexpensive notebooks. Neither of these options fits the needs of someone with communication problems and the use of only one hand. Our goal was to design a device that would allow persons with aphasia and use of one hand to be able to communicate more easily with others on a daily basis. The information gathered from the interviews and observations gave us direction in our design process. We determined that the device should be accessible with one hand, be small in size, have an erasable surface, and have a professional appearance. Our final design is the Fold-A-Board communication device. The Fold-A-Board communication device meets all the requirements for an easy-to-use writing device.

  12. Inverse Kinetics

    2000-03-20

    Given the space-independent, one energy group reactor kinetics equations and the initial conditions, this prgram determines the time variation of reactivity required to produce the given input of flux-time data.

  13. Substrate protein folds while it is bound to the ATP-independent chaperone Spy.

    PubMed

    Stull, Frederick; Koldewey, Philipp; Humes, Julia R; Radford, Sheena E; Bardwell, James C A

    2016-01-01

    Chaperones assist in the folding of many proteins in the cell. Although the most well-studied chaperones use cycles of ATP binding and hydrolysis to assist in protein folding, a number of chaperones have been identified that promote folding in the absence of high-energy cofactors. Precisely how ATP-independent chaperones accomplish this feat is unclear. Here we characterized the kinetic mechanism of substrate folding by the small ATP-independent chaperone Spy from Escherichia coli. Spy rapidly associates with its substrate, immunity protein 7 (Im7), thereby eliminating Im7's potential for aggregation. Remarkably, Spy then allows Im7 to fully fold into its native state while it remains bound to the surface of the chaperone. These results establish a potentially widespread mechanism whereby ATP-independent chaperones assist in protein refolding. They also provide compelling evidence that substrate proteins can fold while being continuously bound to a chaperone. PMID:26619265

  14. Substrate protein folds while it is bound to the ATP-independent chaperone Spy.

    PubMed

    Stull, Frederick; Koldewey, Philipp; Humes, Julia R; Radford, Sheena E; Bardwell, James C A

    2016-01-01

    Chaperones assist in the folding of many proteins in the cell. Although the most well-studied chaperones use cycles of ATP binding and hydrolysis to assist in protein folding, a number of chaperones have been identified that promote folding in the absence of high-energy cofactors. Precisely how ATP-independent chaperones accomplish this feat is unclear. Here we characterized the kinetic mechanism of substrate folding by the small ATP-independent chaperone Spy from Escherichia coli. Spy rapidly associates with its substrate, immunity protein 7 (Im7), thereby eliminating Im7's potential for aggregation. Remarkably, Spy then allows Im7 to fully fold into its native state while it remains bound to the surface of the chaperone. These results establish a potentially widespread mechanism whereby ATP-independent chaperones assist in protein refolding. They also provide compelling evidence that substrate proteins can fold while being continuously bound to a chaperone.

  15. A similar in vitro and in cell lysate folding intermediate for the FF domain.

    PubMed

    Latham, Michael P; Kay, Lewis E

    2014-09-23

    Understanding the mechanisms by which proteins fold into their three-dimensional structures, including a description of the intermediates that are formed during the folding process, remains a goal of protein science. Most studies are performed under carefully controlled conditions in which the folding reaction is monitored in a buffer solution that is far from the natural milieu of the cell. Here, we have used (13)C and (1)H relaxation dispersion NMR spectroscopy to study folding of the FF domain in both Escherichia coli and Saccharomyces cerevisiae cellular lysates. We find that a conformationally excited state is populated in both lysates, which is very similar in structure to a folding intermediate observed in previous studies in buffer, with the kinetics and thermodynamics of the interconversion between native and intermediate conformers somewhat changed. The results point to the importance of extending folding studies beyond the test tube yet emphasize that insights can be obtained through careful experiments recorded in controlled buffer solutions.

  16. A New Heuristic Algorithm for Protein Folding in the HP Model.

    PubMed

    Traykov, Metodi; Angelov, Slav; Yanev, Nicola

    2016-08-01

    This article presents an efficient heuristic for protein folding. The protein folding problem is to predict the compact three-dimensional structure of a protein based on its amino acid sequence. The focus is on an original integer programming model derived from a platform used for Contact Map Overlap problem.

  17. Fast Monte Carlo, slow protein kinetics and perfect loop closure

    NASA Astrophysics Data System (ADS)

    Wedemeyer, William Joseph

    This thesis presents experimental studies of proteins and computational methods which may help in simulations of proteins. The experimental chapters focus on the folding and unfolding of bovine pancreatic ribonuclease A. Methods are developed for tracking the cis-trans isomerization of individual prolines under folding and unfolding conditions, and for identifying critical folding structures by assessing the effects of individual incorrect X-Pro isomers on the conformational folding. The major β-hairpin region is identified as more critical than the C-terminal hydrophobic core. Site- directed mutagenesis of three nearby tyrosines to phenylalanine indicates that tyrosyl hydrogen bonds are essential to rapid conformational folding. Another experimental chapter presents an analytic solution of the kinetics of competitive binding, which is applied to estimating the association and dissociation rate constants of hirudin and thrombin. An extension of this method is proposed to obtain kinetic rate constants for the conformational folding and unfolding of individual parts of a protein. The analytic solution is found to be roughly one-hundred-fold more efficient than the best numerical integrators. The theoretical chapters present methods potentially useful in protein simulations. The loop closure problem is solved geometrically, allowing the protein to be broken into segments which move quasi-independently. Two bootstrap Monte Carlo methods are developed for sampling functions that are characterized by high anisotropy, e.g. long, narrow valleys. Two chapters are devoted to smoothing methods; the first develops a method for exploiting smoothing to evaluate the energy in order N (not N2) time, while the second examines the limitations of one smoothing method, the Diffusion Equation Method, and suggests improvements to its smoothing transformation and reversing procedure. One chapter develops a highly optimized simulation package for lattice heteropolymers by careful choice

  18. Flexibility damps macromolecular crowding effects on protein folding dynamics: Application to the murine prion protein (121-231)

    NASA Astrophysics Data System (ADS)

    Bergasa-Caceres, Fernando; Rabitz, Herschel A.

    2014-01-01

    A model of protein folding kinetics is applied to study the combined effects of protein flexibility and macromolecular crowding on protein folding rate and stability. It is found that the increase in stability and folding rate promoted by macromolecular crowding is damped for proteins with highly flexible native structures. The model is applied to the folding dynamics of the murine prion protein (121-231). It is found that the high flexibility of the native isoform of the murine prion protein (121-231) reduces the effects of macromolecular crowding on its folding dynamics. The relevance of these findings for the pathogenic mechanism are discussed.

  19. Hydrophobic and Ionic-Interactions in Bulk and Confined Water with Implications for Collapse and Folding of Proteins

    NASA Astrophysics Data System (ADS)

    Vaitheeswaran, S.; Chen, Jie; Thirumalai, D.

    2011-10-01

    Water and water-mediated interactions determine the thermodynamics and kinetics of protein folding, protein aggregation and self-assembly in confined spaces. To obtain insights into the role of water in the context of folding problems, we describe computer simulations of a few related model systems. The dynamics of collapse of eicosane shows that upon expulsion of water the linear hydrocarbon chain adopts an ordered helical hairpin structure with 1.5 turns. The structure of dimer of eicosane molecules has two well ordered helical hairpins that are stacked perpendicular to each other. As a prelude to studying folding in confined spaces we used simulations to understand changes in hydrophobic and ionic interactions in nano-sized water droplets. Solvation of hydrophobic and charged species change drastically in nano-scale water droplets. Hydrophobic species are localized at the boundary. The tendency of ions to be at the boundary where water density is low increases as the charge density decreases. The interactions between hydrophobic, polar, and charged residue are also profoundly altered in confined spaces. Using the results of computer simulations and accounting for loss of chain entropy upon confinement we argue and then demonstrate, using simulations in explicit water, that ordered states of generic amphiphilic peptide sequences should be stabilized in cylindrical nanopores.

  20. Oxygen isotope exchange kinetics of mineral pairs in closed and open systems: Applications to problems of hydrothermal alteration of igneous rocks and Precambrian iron formations

    USGS Publications Warehouse

    Gregory, R.T.; Criss, R.E.; Taylor, H.P.

    1989-01-01

    The systematics of stable-isotope exchange between minerals and fluids are examined in the context of modal mineralogical variations and mass-balance considerations, both in closed and in open systems. On mineral-pair ??18O plots, samples from terranes that have exchanged with large amounts of fluid typically map out steep positively-sloped non-equilibrium arrays. Analytical models are derived to explain these effects; these models allow for different exchange rates between the various minerals and the external fluids, as well as different fluid fluxes. The steep arrays are adequately modelled by calculated isochron lines that involve the whole family of possible exchange trajectories. These isochrons have initially-steep near-vertical positive slopes that rotate toward a 45?? equilibrium slope as the exchange process proceeds to completion. The actual data-point array is thus analogous to the hand of an "isotopic clock" that measures the duration of the hydrothermal episode. The dimensionless ratio of the volumetric fluid flux to the kinetic rate parameter ( u k) determines the shape of each individual exchange trajectory. In a fluid-buffered system ( u k ??? 1), the solutions to the equations: (1) are independent of the mole fractions of the solid phases; (2) correspond to Taylor's open-system water/rock equation; and (3) yield straight-line isochrons that have slopes that approach 1 f, where f is the fraction reacted of the more sluggishly exchanging mineral. The isochrons for this simple exchange model are closely congruent with the isochrons calculated for all of the more complex models, thereby simplifying the application of theory to actual hydrothermal systems in nature. In all of the models an order of magnitude of time (in units of kt) separates steep non-equilibrium arrays (e.g., slope ??? 10) from arrays approaching an equilibrium slope of unity on a ??-?? diagram. Because we know the approximate lifetimes of many hydrothermal systems from geologic and

  1. Geometry of Miura-folded metamaterials

    PubMed Central

    Schenk, Mark; Guest, Simon D.

    2013-01-01

    This paper describes two folded metamaterials based on the Miura-ori fold pattern. The structural mechanics of these metamaterials are dominated by the kinematics of the folding, which only depends on the geometry and therefore is scale-independent. First, a folded shell structure is introduced, where the fold pattern provides a negative Poisson’s ratio for in-plane deformations and a positive Poisson’s ratio for out-of-plane bending. Second, a cellular metamaterial is described based on a stacking of individual folded layers, where the folding kinematics are compatible between layers. Additional freedom in the design of the metamaterial can be achieved by varying the fold pattern within each layer. PMID:23401549

  2. Paper Folding for the Mathematics Class.

    ERIC Educational Resources Information Center

    Johnson, Donovan A.

    Directions are given for folding paper to show geometric figures and relationships. Topics covered are folding the basic constructions, geometric concepts concerning triangles and quadrilaterals, circle relationships, products and factors, polygon constructions, symmetry, conic sections, and recreations. (DT)

  3. The parallel universe of RNA folding.

    PubMed

    Batey, R T; Doudna, J A

    1998-05-01

    How do large RNA molecules find their active conformations among a universe of possible structures? Two recent studies reveal that RNA folding is a rapid and ordered process, with surprising similarities to protein folding mechanisms.

  4. Protein Stability, Folding and Misfolding in Human PGK1 Deficiency.

    PubMed

    Valentini, Giovanna; Maggi, Maristella; Pey, Angel L

    2013-01-01

    Conformational diseases are often caused by mutations, altering protein folding and stability in vivo. We review here our recent work on the effects of mutations on the human phosphoglycerate kinase 1 (hPGK1), with a particular focus on thermodynamics and kinetics of protein folding and misfolding. Expression analyses and in vitro biophysical studies indicate that disease-causing mutations enhance protein aggregation propensity. We found a strong correlation among protein aggregation propensity, thermodynamic stability, cooperativity and dynamics. Comparison of folding and unfolding properties with previous reports in PGKs from other species suggests that hPGK1 is very sensitive to mutations leading to enhance protein aggregation through changes in protein folding cooperativity and the structure of the relevant denaturation transition state for aggregation. Overall, we provide a mechanistic framework for protein misfolding of hPGK1, which is insightful to develop new therapeutic strategies aimed to target native state stability and foldability in hPGK1 deficient patients. PMID:24970202

  5. Folding of Small Proteins Using Constrained Molecular Dynamics

    PubMed Central

    Balaraman, Gouthaman S.; Park, In-Hee; Jain, Abhinandan; Vaidehi, Nagarajan

    2011-01-01

    The focus of this paper is to examine whether conformational search using constrained molecular dynamics (MD) method is more enhanced and enriched towards “native-like” structures compared to all-atom MD for the protein folding as a model problem. Constrained MD methods provide an alternate MD tool for protein structure prediction and structure refinement. It is computationally expensive to perform all-atom simulations of protein folding because the processes occur on a timescale of microseconds. Compared to the all-atom MD simulation, constrained MD methods have the advantage that stable dynamics can be achieved for larger time steps and the number of degrees of freedom is an order of magnitude smaller, leading to a decrease in computational cost. We have developed a generalized constrained MD method that allows the user to “freeze and thaw” torsional degrees of freedom as fit for the problem studied. We have used this method to perform all-torsion constrained MD in implicit solvent coupled with the replica exchange method to study folding of small proteins with various secondary structural motifs such as, α-helix (polyalanine, WALP16), β-turn (1E0Q), and a mixed motif protein (Trp-cage). We demonstrate that constrained MD replica exchange method exhibits a wider conformational search than all-atom MD with increased enrichment of near native structures. “Hierarchical” constrained MD simulations, where the partially formed helical regions in the initial stretch of the all-torsion folding simulation trajectory of Trp-cage were frozen, showed a better sampling of near native structures than all-torsion constrained MD simulations. This is in agreement with the zipping-and-assembly folding model put forth by Dill and coworkers for folding proteins. The use of hierarchical “freeze and thaw” clustering schemes in constrained MD simulation can be used to sample conformations that contribute significantly to folding of proteins. PMID:21591767

  6. Solitons and protein folding: An In Silico experiment

    NASA Astrophysics Data System (ADS)

    Ilieva, N.; Dai, J.; Sieradzan, A.; Niemi, A.

    2015-10-01

    Protein folding [1] is the process of formation of a functional 3D structure from a random coil — the shape in which amino-acid chains leave the ribosome. Anfinsen's dogma states that the native 3D shape of a protein is completely determined by protein's amino acid sequence. Despite the progress in understanding the process rate and the success in folding prediction for some small proteins, with presently available physics-based methods it is not yet possible to reliably deduce the shape of a biologically active protein from its amino acid sequence. The protein-folding problem endures as one of the most important unresolved problems in science; it addresses the origin of life itself. Furthermore, a wrong fold is a common cause for a protein to lose its function or even endanger the living organism. Soliton solutions of a generalized discrete non-linear Schrödinger equation (GDNLSE) obtained from the energy function in terms of bond and torsion angles κ and τ provide a constructive theoretical framework for describing protein folds and folding patterns [2]. Here we study the dynamics of this process by means of molecular-dynamics simulations. The soliton manifestation is the pattern helix-loop-helix in the secondary structure of the protein, which explains the importance of understanding loop formation in helical proteins. We performed in silico experiments for unfolding one subunit of the core structure of gp41 from the HIV envelope glycoprotein (PDB ID: 1AIK [3]) by molecular-dynamics simulations with the MD package GROMACS. We analyzed 80 ns trajectories, obtained with one united-atom and two different all-atom force fields, to justify the side-chain orientation quantification scheme adopted in the studies and to eliminate force-field based artifacts. Our results are compatible with the soliton model of protein folding and provide first insight into soliton-formation dynamics.

  7. Solitons and protein folding: An In Silico experiment

    SciTech Connect

    Ilieva, N.; Dai, J.; Sieradzan, A.; Niemi, A.

    2015-10-28

    Protein folding [1] is the process of formation of a functional 3D structure from a random coil — the shape in which amino-acid chains leave the ribosome. Anfinsen’s dogma states that the native 3D shape of a protein is completely determined by protein’s amino acid sequence. Despite the progress in understanding the process rate and the success in folding prediction for some small proteins, with presently available physics-based methods it is not yet possible to reliably deduce the shape of a biologically active protein from its amino acid sequence. The protein-folding problem endures as one of the most important unresolved problems in science; it addresses the origin of life itself. Furthermore, a wrong fold is a common cause for a protein to lose its function or even endanger the living organism. Soliton solutions of a generalized discrete non-linear Schrödinger equation (GDNLSE) obtained from the energy function in terms of bond and torsion angles κ and τ provide a constructive theoretical framework for describing protein folds and folding patterns [2]. Here we study the dynamics of this process by means of molecular-dynamics simulations. The soliton manifestation is the pattern helix–loop–helix in the secondary structure of the protein, which explains the importance of understanding loop formation in helical proteins. We performed in silico experiments for unfolding one subunit of the core structure of gp41 from the HIV envelope glycoprotein (PDB ID: 1AIK [3]) by molecular-dynamics simulations with the MD package GROMACS. We analyzed 80 ns trajectories, obtained with one united-atom and two different all-atom force fields, to justify the side-chain orientation quantification scheme adopted in the studies and to eliminate force-field based artifacts. Our results are compatible with the soliton model of protein folding and provide first insight into soliton-formation dynamics.

  8. Mapping fast protein folding with multiple-site fluorescent probes

    PubMed Central

    Prigozhin, Maxim B.; Chao, Shu-Han; Sukenik, Shahar; Pogorelov, Taras V.; Gruebele, Martin

    2015-01-01

    Fast protein folding involves complex dynamics in many degrees of freedom, yet microsecond folding experiments provide only low-resolution structural information. We enhance the structural resolution of the five-helix bundle protein λ6–85 by engineering into it three fluorescent tryptophan–tyrosine contact probes. The probes report on distances between three different helix pairs: 1–2, 1–3, and 3–2. Temperature jump relaxation experiments on these three mutants reveal two different kinetic timescales: a slower timescale for 1–3 and a faster one for the two contacts involving helix 2. We hypothesize that these differences arise from a single folding mechanism that forms contacts on different timescales, and not from changes of mechanism due to adding the probes. To test this hypothesis, we analyzed the corresponding three distances in one published single-trajectory all-atom molecular-dynamics simulation of a similar mutant. Autocorrelation analysis of the trajectory reveals the same “slow” and “fast” distance change as does experiment, but on a faster timescale; smoothing the trajectory in time shows that this ordering is robust and persists into the microsecond folding timescale. Structural investigation of the all-atom computational data suggests that helix 2 misfolds to produce a short-lived off-pathway trap, in agreement with the experimental finding that the 1–2 and 3–2 distances involving helix 2 contacts form a kinetic grouping distinct from 1 to 3. Our work demonstrates that comparison between experiment and simulation can be extended to several order parameters, providing a stronger mechanistic test. PMID:26080403

  9. Mapping fast protein folding with multiple-site fluorescent probes.

    PubMed

    Prigozhin, Maxim B; Chao, Shu-Han; Sukenik, Shahar; Pogorelov, Taras V; Gruebele, Martin

    2015-06-30

    Fast protein folding involves complex dynamics in many degrees of freedom, yet microsecond folding experiments provide only low-resolution structural information. We enhance the structural resolution of the five-helix bundle protein λ6-85 by engineering into it three fluorescent tryptophan-tyrosine contact probes. The probes report on distances between three different helix pairs: 1-2, 1-3, and 3-2. Temperature jump relaxation experiments on these three mutants reveal two different kinetic timescales: a slower timescale for 1-3 and a faster one for the two contacts involving helix 2. We hypothesize that these differences arise from a single folding mechanism that forms contacts on different timescales, and not from changes of mechanism due to adding the probes. To test this hypothesis, we analyzed the corresponding three distances in one published single-trajectory all-atom molecular-dynamics simulation of a similar mutant. Autocorrelation analysis of the trajectory reveals the same "slow" and "fast" distance change as does experiment, but on a faster timescale; smoothing the trajectory in time shows that this ordering is robust and persists into the microsecond folding timescale. Structural investigation of the all-atom computational data suggests that helix 2 misfolds to produce a short-lived off-pathway trap, in agreement with the experimental finding that the 1-2 and 3-2 distances involving helix 2 contacts form a kinetic grouping distinct from 1 to 3. Our work demonstrates that comparison between experiment and simulation can be extended to several order parameters, providing a stronger mechanistic test. PMID:26080403

  10. Extracting Information from Folds in Rocks.

    ERIC Educational Resources Information Center

    Hudleston, Peter John

    1986-01-01

    Describes the three processes of folding in rocks: buckling, bending, and passive folding. Discusses how geometrical properties and strain distributions help to identify which processes produce natural folds, and also provides information about the mechanical properties of rocks, and the sense of shear in shear zones. (TW)

  11. Dynamics of Folds in the Plane

    ERIC Educational Resources Information Center

    Krylov, Nikolai A.; Rogers, Edwin L.

    2011-01-01

    Take a strip of paper and fold a crease intersecting the long edges, creating two angles. Choose one edge and consider the angle with the crease. Fold the opposite edge along the crease, creating a new crease that bisects the angle. Fold again, this time using the newly created crease and the initial edge, creating a new angle along the chosen…

  12. Surgical revision of the upper eyelid fold.

    PubMed

    Cies, W A; Baylis, H I

    1975-12-01

    We performed surgery on 107 patients primarily with blepharoptosis and eyelid fold abnormalities, between 1973 and 1974. Production of an eyelid fold at the time of an initial blepharoptosis procedure should be a primary goal. Lack of a distinct symmetrical upper eyelid fold constituted a cosmetic blemish and necessitated revision.

  13. Numerical study of human vocal folds vibration using Immersed Finite Element Method

    NASA Astrophysics Data System (ADS)

    Wang, Xingshi; Zhang, Lucy; Krane, Michael

    2011-11-01

    The voice production procedure is a self-oscillating, fluid-structure interaction problem. In this study, the vocal folds vibration during phonation will be simulated by self-oscillated layered-structure vocal folds model, using Immersed Finite Element Method. With the numerical results, we will find out the vocal folds vibration pattern, and also show how the lung pressure, stiffness and geometry of vocal folds will affect the vocal folds vibration. With further analysis, we shall get better understanding of the dynamics of voice production. National Institute on Deafness and Other Communication Disorders.

  14. Deciphering the Structural Basis That Guides the Oxidative Folding of Leech-derived Tryptase Inhibitor*

    PubMed Central

    Pantoja-Uceda, David; Arolas, Joan L.; Aviles, Francesc X.; Santoro, Jorge; Ventura, Salvador; Sommerhoff, Christian P.

    2009-01-01

    Protein folding mechanisms have remained elusive mainly because of the transient nature of intermediates. Leech-derived tryptase inhibitor (LDTI) is a Kazal-type serine proteinase inhibitor that is emerging as an attractive model for folding studies. It comprises 46 amino acid residues with three disulfide bonds, with one located inside a small triple-stranded antiparallel β-sheet and with two involved in a cystine-stabilized α-helix, a motif that is widely distributed in bioactive peptides. Here, we analyzed the oxidative folding and reductive unfolding of LDTI by chromatographic and disulfide analyses of acid-trapped intermediates. It folds and unfolds, respectively, via sequential oxidation and reduction of the cysteine residues that give rise to a few 1- and 2-disulfide intermediates. Species containing two native disulfide bonds predominate during LDTI folding (IIa and IIc) and unfolding (IIa and IIb). Stop/go folding experiments demonstrate that only intermediate IIa is productive and oxidizes directly into the native form. The NMR structures of acid-trapped and further isolated IIa, IIb, and IIc reveal global folds similar to that of the native protein, including a native-like canonical inhibitory loop. Enzyme kinetics shows that both IIa and IIc are inhibitory-active, which may substantially reduce proteolysis of LDTI during its folding process. The results reported show that the kinetics of the folding reaction is modulated by the specific structural properties of the intermediates and together provide insights into the interdependence of conformational folding and the assembly of native disulfides during oxidative folding. PMID:19820233

  15. Understanding Protein Non-Folding

    PubMed Central

    Uversky, Vladimir N.; Dunker, A. Keith

    2010-01-01

    This review describes the family of intrinsically disordered proteins, members of which fail to form rigid 3-D structures under physiological conditions, either along their entire lengths or only in localized regions. Instead, these intriguing proteins/regions exist as dynamic ensembles within which atom positions and backbone Ramachandran angles exhibit extreme temporal fluctuations without specific equilibrium values. Many of these intrinsically disordered proteins are known to carry out important biological functions which, in fact, depend on the absence of specific 3-D structure. The existence of such proteins does not fit the prevailing structure-function paradigm, which states that unique 3-D structure is a prerequisite to function. Thus, the protein structure-function paradigm has to be expanded to include intrinsically disordered proteins and alternative relationships among protein sequence, structure, and function. This shift in the paradigm represents a major breakthrough for biochemistry, biophysics and molecular biology, as it opens new levels of understanding with regard to the complex life of proteins. This review will try to answer the following questions: How were intrinsically disordered proteins discovered? Why don't these proteins fold? What is so special about intrinsic disorder? What are the functional advantages of disordered proteins/regions? What is the functional repertoire of these proteins? What are the relationships between intrinsically disordered proteins and human diseases? PMID:20117254

  16. Chemical Kinetics Laboratory Discussion Worksheet.

    PubMed

    Demoin, Dustin Wayne; Jurisson, Silvia S

    2013-09-10

    A laboratory discussion worksheet and its answer key provide instructors and students a discussion model to further the students' understanding of chemical kinetics. This discussion worksheet includes a section for students to augment their previous knowledge about chemical kinetics measurements, an initial check on students' understanding of basic concepts, a group participation model where students work on solving complex-conceptual problems, and a conclusion to help students connect this discussion to their laboratory or lecture class. Additionally, the worksheet has a detailed solution to a more advanced problem to help students understand how the concepts they have put together relate to problems they will encounter during later formal assessments.

  17. Anatomy and Histology of an Epicanthal Fold.

    PubMed

    Park, Jae Woo; Hwang, Kun

    2016-06-01

    The aim of this study is to elucidate the precise anatomical and histological detail of the epicanthal fold.Thirty-two hemifaces of 16 Korean adult cadavers were used in this study (30 hemifaces with an epicanthal fold, 2 without an epicanthal fold). In 2 patients who had an epicanthoplasty, the epicanthal folds were sampled.In a dissection, the periorbital skin and subcutaneous tissues were removed and the epicanthal fold was observed in relation to each part of the orbicularis oculi muscle. Specimens including the epicanthal fold were embeddedin in paraffin, sectioned at 10 um, and stained with Hematoxylin-Eosin. The horizontal section in the level of the paplebral fissure was made and the prepared slides were observed under a light microscope.In the specimens without an epicanthal fold, no connection between the upper preseptal muscle and the lower preseptal muscle was found. In the specimens with an epicanthal fold, a connection of the upper preseptal muscle to the lower preseptal muscle was observed. It was present in all 15 hemifaces (100%). There was no connection between the pretarsal muscles. In a horizontal section, the epicanthal fold was composed of 3 compartments: an outer skin lining, a core structure, and an innerskin lining. The core structure was mainly composed of muscular fibers and fibrotic tissue and they were intermingled.Surgeons should be aware of the anatomical details of an epicanthal fold. In removing or reconstructing an epicanthal fold, the fibromuscular core band should also be removed or reconstructed.

  18. Geomorphology and fold growth, southern Tunisia.

    NASA Astrophysics Data System (ADS)

    Ahamadi, R.; Mercier, E.; Ouali, J.; van Vliet-Lanoë, B.; Mansy, J. L.; Launeau, P.; Rekis, F.

    2003-04-01

    At the northern edge of the Gafsa basin, a fold belt, the Metlaoui chain mostly rised from the Upper Miocene till today [1]. This region, endoreic since the Oligocene, is for lithologic and climate reasons a good laboratory to test various theories concerning fold growth. The origin of the fold belt is link to a main detachment level at the base of the Jurassic series (seismic profiles). The folding is facilitate with few strong calcareous beds by the dominance of soft lithologic series. This enhances the sliding along secondary detachment levels. Geomorphological, hydrological and chronological arguments prove that the folds mostly deformed as a "fault propagation fold" [2]. Furthermore, detailed field observations validate several features first observed by modelling of "fault propagation fold": 1) the stability of the roof of the fold, 2) the propagation of a knock-fold in front of the fold, disturbing the shape of pediments [3] and 3) the backward migration and extent of the backslope of the fold, in association with a backward migration of the depot centre of the syntectonic sedimentation. [1] Outtani, F., B. Addoum, E. Mercier, D. Frizon de Lamotte, J. Andrieux, Tectonophysics, 249, 233-248, 1995. [2] Suppe, J., and D.A. Medwedeff, , Eclogae geol. Helv., 83(3), 409-454, 1990. [3] Rafini S. and E. Mercier (2002). Sedimentary Geology, 146, 75-89

  19. Metal ion specificities for folding and cleavage activity in the Schistosoma hammerhead ribozyme

    PubMed Central

    Boots, Jennifer L.; Canny, Marella D.; Azimi, Ehsan; Pardi, Arthur

    2008-01-01

    The effects of various metal ions on cleavage activity and global folding have been studied in the extended Schistosoma hammerhead ribozyme. Fluorescence resonance energy transfer was used to probe global folding as a function of various monovalent and divalent metal ions in this ribozyme. The divalent metals ions Ca2+, Mg2+, Mn2+, and Sr2+ have a relatively small variation (less than sixfold) in their ability to globally fold the hammerhead ribozyme, which contrasts with the very large difference (>10,000-fold) in apparent rate constants for cleavage for these divalent metal ions in single-turnover kinetic experiments. There is still a very large range (>4600-fold) in the apparent rate constants for cleavage for these divalent metal ions measured in high salt (2 M NaCl) conditions where the ribozyme is globally folded. These results demonstrate that the identity of the divalent metal ion has little effect on global folding of the Schistosoma hammerhead ribozyme, whereas it has a very large effect on the cleavage kinetics. Mechanisms by which the identity of the divalent metal ion can have such a large effect on cleavage activity in the Schistosoma hammerhead ribozyme are discussed. PMID:18755844

  20. Functional analysis of propeptide as an intramolecular chaperone for in vivo folding of subtilisin nattokinase.

    PubMed

    Jia, Yan; Liu, Hui; Bao, Wei; Weng, Meizhi; Chen, Wei; Cai, Yongjun; Zheng, Zhongliang; Zou, Guolin

    2010-12-01

    Here, we show that during in vivo folding of the precursor, the propeptide of subtilisin nattokinase functions as an intramolecular chaperone (IMC) that organises the in vivo folding of the subtilisin domain. Two residues belonging to β-strands formed by conserved regions of the IMC are crucial for the folding of the subtilisin domain through direct interactions. An identical protease can fold into different conformations in vivo due to the action of a mutated IMC, resulting in different kinetic parameters. Some interfacial changes involving conserved regions, even those induced by the subtilisin domain, blocked subtilisin folding and altered its conformation. Insight into the interaction between the subtilisin and IMC domains is provided by a three-dimensional structural model.

  1. Mechanisms of protein-folding diseases at a glance

    PubMed Central

    Valastyan, Julie S.; Lindquist, Susan

    2014-01-01

    For a protein to function appropriately, it must first achieve its proper conformation and location within the crowded environment inside the cell. Multiple chaperone systems are required to fold proteins correctly. In addition, degradation pathways participate by destroying improperly folded proteins. The intricacy of this multisystem process provides many opportunities for error. Furthermore, mutations cause misfolded, nonfunctional forms of proteins to accumulate. As a result, many pathological conditions are fundamentally rooted in the protein-folding problem that all cells must solve to maintain their function and integrity. Here, to illustrate the breadth of this phenomenon, we describe five examples of protein-misfolding events that can lead to disease: improper degradation, mislocalization, dominant-negative mutations, structural alterations that establish novel toxic functions, and amyloid accumulation. In each case, we will highlight current therapeutic options for battling such diseases. PMID:24396149

  2. Stress and strain evolution of folding rocks

    NASA Astrophysics Data System (ADS)

    Llorens, Maria-Gema; Griera, Albert; Bons, Paul; Gomez-Rivas, Enrique; Weikusat, Ilka

    2015-04-01

    One of the main objectives of structural geology is to unravel rock deformation histories. Fold shapes can be used to estimate the orientation and amount of strain associated with folding. However, much more information on rheology and kinematics can potentially be extracted from fold geometries (Llorens et al., 2013a). We can study the development of folds, quantify the relationships between the different parameters that determine their geometries and estimate their mechanical evolution. This approach allows us to better understand and predict not only rock but also ice deformation. One of the main parameters in fold development is the viscosity contrast between the folding layer and the matrix in which it is embedded (m), since it determines the initial fold wavelength and the amplification rate of the developing folds. Moreover, non-linear viscous rheology influences fold geometry too (Llorens et al., 2013b). We present a series of 2-dimensional simulations of folding of viscous single layers in pure and simple shear. We vary different parameters in order to compare and determine their influence on the resulting fold patterns and the associated mechanical response of the material. To perform these simulations we use the software platform ELLE (www.elle.ws) with the non-linear viscous finite element code BASIL. The results show that layers thicken at the beginning of deformation in all simulations, and visible folds start earlier or later depending on the viscosity contrast. When folds start to nucleate the layer maximum shear strain decreases, moving away from the theoretical trend for homogeneous strain (no folding). This allows the accurate determination of the onset of folding. Maximum deviatoric stresses are higher in power-law than in linear-viscosity materials, and it is initially double in pure shear than in simple shear conditions. Therefore, folding a competent layer requires less work in simple than in pure shear. The maximum deviatoric stress

  3. Some aspects of vocal fold bowing.

    PubMed

    Tanaka, S; Hirano, M; Chijiwa, K

    1994-05-01

    Bowing of the vocal fold frequently occurs in patients with vocal fold paralysis (VFP), those with sulcus vocalis, and those who have had laser surgery. Additionally, there are vocal folds that present bowing with no noticeable organic lesion. For the purpose of investigating the causes and mechanisms of vocal fold bowing, consecutive fiberscopic videorecordings of 127 patients with VFP, 33 with sulcus vocalis, 33 with laser surgery, and 33 with dysphonia having no clinically noticeable organic lesion were reviewed. Sixty-nine percent of the paralyzed vocal folds had bowing, and the occurrence of bowing was significantly related to the activity of the thyroarytenoid muscle as measured by electromyography. The cricothyroid activity had no significant relationship to vocal fold bowing. All vocal folds with sulcus presented with bowing. Thirty-five percent of the vocal folds that had had laser surgery had bowing. The extent of tissue removal was closely related to the occurrence of bowing. Twelve cases with no organic lesion had vocal fold bowing. Of these 12 patients, 8 were male and 9 were older than 60 years. Some aging process in the mucosa was presumed to be the cause of the bowing in this age group of patients without clinically noticeable organic lesions. Causes of vocal fold bowing in the younger group of patients without organic lesions were not determined in this study.

  4. Some aspects of vocal fold bowing.

    PubMed

    Tanaka, S; Hirano, M; Chijiwa, K

    1994-05-01

    Bowing of the vocal fold frequently occurs in patients with vocal fold paralysis (VFP), those with sulcus vocalis, and those who have had laser surgery. Additionally, there are vocal folds that present bowing with no noticeable organic lesion. For the purpose of investigating the causes and mechanisms of vocal fold bowing, consecutive fiberscopic videorecordings of 127 patients with VFP, 33 with sulcus vocalis, 33 with laser surgery, and 33 with dysphonia having no clinically noticeable organic lesion were reviewed. Sixty-nine percent of the paralyzed vocal folds had bowing, and the occurrence of bowing was significantly related to the activity of the thyroarytenoid muscle as measured by electromyography. The cricothyroid activity had no significant relationship to vocal fold bowing. All vocal folds with sulcus presented with bowing. Thirty-five percent of the vocal folds that had had laser surgery had bowing. The extent of tissue removal was closely related to the occurrence of bowing. Twelve cases with no organic lesion had vocal fold bowing. Of these 12 patients, 8 were male and 9 were older than 60 years. Some aging process in the mucosa was presumed to be the cause of the bowing in this age group of patients without clinically noticeable organic lesions. Causes of vocal fold bowing in the younger group of patients without organic lesions were not determined in this study. PMID:8179251

  5. Folding of synthetic homogeneous glycoproteins in the presence of a glycoprotein folding sensor enzyme.

    PubMed

    Dedola, Simone; Izumi, Masayuki; Makimura, Yutaka; Seko, Akira; Kanamori, Akiko; Sakono, Masafumi; Ito, Yukishige; Kajihara, Yasuhiro

    2014-03-10

    UDP-glucose:glycoprotein glucosyltransferase (UGGT) plays a key role in recognizing folded and misfolded glycoproteins in the glycoprotein quality control system of the endoplasmic reticulum. UGGT detects misfolded glycoproteins and re-glucosylates them as a tag for misfolded glycoproteins. A flexible model to reproduce in vitro folding of a glycoprotein in the presence of UGGT in a mixture containing correctly folded, folding intermediates, and misfolded glycoproteins is described. The data demonstrates that UGGT can re-glucosylate all intermediates in the in vitro folding experiments, thus indicating that UGGT inspects not only final folded products, but also the glycoprotein folding intermediates.

  6. Protein folding at atomic resolution: analysis of autonomously folding supersecondary structure motifs by nuclear magnetic resonance.

    PubMed

    Sborgi, Lorenzo; Verma, Abhinav; Sadqi, Mourad; de Alba, Eva; Muñoz, Victor

    2013-01-01

    The study of protein folding has been conventionally hampered by the assumption that all single-domain proteins fold by an all-or-none process (two-state folding) that makes it impossible to resolve folding mechanisms experimentally. Here we describe an experimental method for the thermodynamic analysis of protein folding at atomic resolution using nuclear magnetic resonance (NMR). The method is specifically developed for the study of small proteins that fold autonomously into basic supersecondary structure motifs, and that do so in the sub-millisecond timescale (folding archetypes). From the NMR experiments we obtain hundreds of atomic unfolding curves that are subsequently analyzed leading to the determination of the characteristic network of folding interactions. The application of this approach to a comprehensive catalog of elementary folding archetypes holds the promise of becoming the first experimental approach capable of unraveling the basic rules connecting protein structure and folding mechanism. PMID:22987355

  7. Folding Dynamics of an α Helix and a β Hairpin

    NASA Astrophysics Data System (ADS)

    Hofrichter, James

    1998-03-01

    What processes limit the rate at which proteins fold? In an effort to address this question we have begun to study the dynamics of the formation of loops, α helices and the minimal β structural element, a β hairpin, which must occur on the pathway from random coils to folded proteins. Because these processes occur on time scales of 10-5-10-9 seconds and experimental access to these time scales has been limited, the kinetics of these processes have not been extensively studied. The expectation is that a more complete understanding of the dynamics of these microprocesses will provide constraints on possible mechanisms for the overall folding of more complex structures. We have explored the kinetics of the helix-coil transition of a synthetic, 21-residue peptide: Ac-WAAAH^+(AAARA)_3A-NH2 and of the folding of a 16 residue β hairpin from protein G B1 using the nanosecond temperature jump technique. Both processes were studied by monitoring tryptophan fluorescence. In the helical peptide, the quantum yield of tryptophan decreases as a result of the interaction between tryptophan in position 1 with the protonated histidine in position 5. In the native conformation of the hairpin, it increases because it forms part of a hydrophobic cluster which stabilizes the native conformation (in a peptide in which a dansylated lysine is incorporated at the C-terminus the fluorescence is quenched). At 300 K, the relaxation time for the helix-coil transition is ~ 250 ns and that for the hairpin-coil transition is ~ 2.2 μs, about 10 times slower. The apparent activation energies are 6.8 kcal/mol for the helix and 10 kcal/mol for the hairpin. We have developed simple kinetic models for these processes which incorporate the sequence- and position-dependent properties known from equilibrium studies and the single-sequence approximation. These models provide a remarkably consistent picture of the dynamics, permitting us to extract information on both the microscopic rates for the c

  8. On the universe of protein folds.

    PubMed

    Kolodny, Rachel; Pereyaslavets, Leonid; Samson, Abraham O; Levitt, Michael

    2013-01-01

    In the fifty years since the first atomic structure of a protein was revealed, tens of thousands of additional structures have been solved. Like all objects in biology, proteins structures show common patterns that seem to define family relationships. Classification of proteins structures, which started in the 1970s with about a dozen structures, has continued with increasing enthusiasm, leading to two main fold classifications, SCOP and CATH, as well as many additional databases. Classification is complicated by deciding what constitutes a domain, the fundamental unit of structure. Also difficult is deciding when two given structures are similar. Like all of biology, fold classification is beset by exceptions to all rules. Thus, the perspectives of protein fold space that the fold classifications offer differ from each other. In spite of these ambiguities, fold classifications are useful for prediction of structure and function. Studying the characteristics of fold space can shed light on protein evolution and the physical laws that govern protein behavior.

  9. Protein Vivisection Reveals Elusive Intermediates in Folding

    SciTech Connect

    Zheng, Zhongzhou; Sosnick, Tobin R.

    2010-05-25

    Although most folding intermediates escape detection, their characterization is crucial to the elucidation of folding mechanisms. Here, we outline a powerful strategy to populate partially unfolded intermediates: A buried aliphatic residue is substituted with a charged residue (e.g., Leu {yields} Glu{sup -}) to destabilize and unfold a specific region of the protein. We applied this strategy to ubiquitin, reversibly trapping a folding intermediate in which the {beta}5-strand is unfolded. The intermediate refolds to a native-like structure upon charge neutralization under mildly acidic conditions. Characterization of the trapped intermediate using NMR and hydrogen exchange methods identifies a second folding intermediate and reveals the order and free energies of the two major folding events on the native side of the rate-limiting step. This general strategy may be combined with other methods and have broad applications in the study of protein folding and other reactions that require trapping of high-energy states.

  10. Protein vivisection reveals elusive intermediates in folding.

    PubMed

    Zheng, Zhongzhou; Sosnick, Tobin R

    2010-04-01

    Although most folding intermediates escape detection, their characterization is crucial to the elucidation of folding mechanisms. Here, we outline a powerful strategy to populate partially unfolded intermediates: A buried aliphatic residue is substituted with a charged residue (e.g., Leu-->Glu(-)) to destabilize and unfold a specific region of the protein. We applied this strategy to ubiquitin, reversibly trapping a folding intermediate in which the beta5-strand is unfolded. The intermediate refolds to a native-like structure upon charge neutralization under mildly acidic conditions. Characterization of the trapped intermediate using NMR and hydrogen exchange methods identifies a second folding intermediate and reveals the order and free energies of the two major folding events on the native side of the rate-limiting step. This general strategy may be combined with other methods and have broad applications in the study of protein folding and other reactions that require trapping of high-energy states.

  11. Statistical mechanics of simple models of protein folding and design.

    PubMed Central

    Pande, V S; Grosberg, A Y; Tanaka, T

    1997-01-01

    It is now believed that the primary equilibrium aspects of simple models of protein folding are understood theoretically. However, current theories often resort to rather heavy mathematics to overcome some technical difficulties inherent in the problem or start from a phenomenological model. To this end, we take a new approach in this pedagogical review of the statistical mechanics of protein folding. The benefit of our approach is a drastic mathematical simplification of the theory, without resort to any new approximations or phenomenological prescriptions. Indeed, the results we obtain agree precisely with previous calculations. Because of this simplification, we are able to present here a thorough and self contained treatment of the problem. Topics discussed include the statistical mechanics of the random energy model (REM), tests of the validity of REM as a model for heteropolymer freezing, freezing transition of random sequences, phase diagram of designed ("minimally frustrated") sequences, and the degree to which errors in the interactions employed in simulations of either folding and design can still lead to correct folding behavior. Images FIGURE 2 FIGURE 3 FIGURE 4 FIGURE 6 PMID:9414231

  12. Folding and Stabilization of Native-Sequence-Reversed Proteins.

    PubMed

    Zhang, Yuanzhao; Weber, Jeffrey K; Zhou, Ruhong

    2016-04-26

    Though the problem of sequence-reversed protein folding is largely unexplored, one might speculate that reversed native protein sequences should be significantly more foldable than purely random heteropolymer sequences. In this article, we investigate how the reverse-sequences of native proteins might fold by examining a series of small proteins of increasing structural complexity (α-helix, β-hairpin, α-helix bundle, and α/β-protein). Employing a tandem protein structure prediction algorithmic and molecular dynamics simulation approach, we find that the ability of reverse sequences to adopt native-like folds is strongly influenced by protein size and the flexibility of the native hydrophobic core. For β-hairpins with reverse-sequences that fail to fold, we employ a simple mutational strategy for guiding stable hairpin formation that involves the insertion of amino acids into the β-turn region. This systematic look at reverse sequence duality sheds new light on the problem of protein sequence-structure mapping and may serve to inspire new protein design and protein structure prediction protocols.

  13. Folding and Stabilization of Native-Sequence-Reversed Proteins

    PubMed Central

    Zhang, Yuanzhao; Weber, Jeffrey K; Zhou, Ruhong

    2016-01-01

    Though the problem of sequence-reversed protein folding is largely unexplored, one might speculate that reversed native protein sequences should be significantly more foldable than purely random heteropolymer sequences. In this article, we investigate how the reverse-sequences of native proteins might fold by examining a series of small proteins of increasing structural complexity (α-helix, β-hairpin, α-helix bundle, and α/β-protein). Employing a tandem protein structure prediction algorithmic and molecular dynamics simulation approach, we find that the ability of reverse sequences to adopt native-like folds is strongly influenced by protein size and the flexibility of the native hydrophobic core. For β-hairpins with reverse-sequences that fail to fold, we employ a simple mutational strategy for guiding stable hairpin formation that involves the insertion of amino acids into the β-turn region. This systematic look at reverse sequence duality sheds new light on the problem of protein sequence-structure mapping and may serve to inspire new protein design and protein structure prediction protocols. PMID:27113844

  14. Acquired retinal folds in the cat.

    PubMed

    MacMillan, A D

    1976-06-01

    Retinal folds were found in 5 cats. The apparent cause of the folding was varied: in 1 cat the folds appeared after a localized retinal detachment; in 2 cats the condition accompanied other intraocular abnormalities associated with feline infectious peritonitis; 1 cat had active keratitis, and the retinal changes were thought to have been injury related; and 1 cat, bilaterally affected, had chronic glomerulonephritis. PMID:945253

  15. Protein Folding Mechanism of the Dimeric AmphiphysinII/Bin1 N-BAR Domain

    PubMed Central

    Gruber, Tobias; Balbach, Jochen

    2015-01-01

    The human AmphyphisinII/Bin1 N-BAR domain belongs to the BAR domain superfamily, whose members sense and generate membrane curvatures. The N-BAR domain is a 57 kDa homodimeric protein comprising a six helix bundle. Here we report the protein folding mechanism of this protein as a representative of this protein superfamily. The concentration dependent thermodynamic stability was studied by urea equilibrium transition curves followed by fluorescence and far-UV CD spectroscopy. Kinetic unfolding and refolding experiments, including rapid double and triple mixing techniques, allowed to unravel the complex folding behavior of N-BAR. The equilibrium unfolding transition curve can be described by a two-state process, while the folding kinetics show four refolding phases, an additional burst reaction and two unfolding phases. All fast refolding phases show a rollover in the chevron plot but only one of these phases depends on the protein concentration reporting the dimerization step. Secondary structure formation occurs during the three fast refolding phases. The slowest phase can be assigned to a proline isomerization. All kinetic experiments were also followed by fluorescence anisotropy detection to verify the assignment of the dimerization step to the respective folding phase. Based on these experiments we propose for N-BAR two parallel folding pathways towards the homodimeric native state depending on the proline conformation in the unfolded state. PMID:26368922

  16. Outer membrane β-barrel protein folding is physically controlled by periplasmic lipid head groups and BamA.

    PubMed

    Gessmann, Dennis; Chung, Yong Hee; Danoff, Emily J; Plummer, Ashlee M; Sandlin, Clifford W; Zaccai, Nathan R; Fleming, Karen G

    2014-04-22

    Outer membrane β-barrel proteins (OMPs) are crucial for numerous cellular processes in prokaryotes and eukaryotes. Despite extensive studies on OMP biogenesis, it is unclear why OMPs require assembly machineries to fold into their native outer membranes, as they are capable of folding quickly and efficiently through an intrinsic folding pathway in vitro. By investigating the folding of several bacterial OMPs using membranes with naturally occurring Escherichia coli lipids, we show that phosphoethanolamine and phosphoglycerol head groups impose a kinetic barrier to OMP folding. The kinetic retardation of OMP folding places a strong negative pressure against spontaneous incorporation of OMPs into inner bacterial membranes, which would dissipate the proton motive force and undoubtedly kill bacteria. We further show that prefolded β-barrel assembly machinery subunit A (BamA), the evolutionarily conserved, central subunit of the BAM complex, accelerates OMP folding by lowering the kinetic barrier imposed by phosphoethanolamine head groups. Our results suggest that OMP assembly machineries are required in vivo to enable physical control over the spontaneously occurring OMP folding reaction in the periplasm. Mechanistic studies further allowed us to derive a model for BamA function, which explains how OMP assembly can be conserved between prokaryotes and eukaryotes.

  17. Outer membrane β-barrel protein folding is physically controlled by periplasmic lipid head groups and BamA

    PubMed Central

    Gessmann, Dennis; Chung, Yong Hee; Danoff, Emily J.; Plummer, Ashlee M.; Sandlin, Clifford W.; Zaccai, Nathan R.; Fleming, Karen G.

    2014-01-01

    Outer membrane β-barrel proteins (OMPs) are crucial for numerous cellular processes in prokaryotes and eukaryotes. Despite extensive studies on OMP biogenesis, it is unclear why OMPs require assembly machineries to fold into their native outer membranes, as they are capable of folding quickly and efficiently through an intrinsic folding pathway in vitro. By investigating the folding of several bacterial OMPs using membranes with naturally occurring Escherichia coli lipids, we show that phosphoethanolamine and phosphoglycerol head groups impose a kinetic barrier to OMP folding. The kinetic retardation of OMP folding places a strong negative pressure against spontaneous incorporation of OMPs into inner bacterial membranes, which would dissipate the proton motive force and undoubtedly kill bacteria. We further show that prefolded β-barrel assembly machinery subunit A (BamA), the evolutionarily conserved, central subunit of the BAM complex, accelerates OMP folding by lowering the kinetic barrier imposed by phosphoethanolamine head groups. Our results suggest that OMP assembly machineries are required in vivo to enable physical control over the spontaneously occurring OMP folding reaction in the periplasm. Mechanistic studies further allowed us to derive a model for BamA function, which explains how OMP assembly can be conserved between prokaryotes and eukaryotes. PMID:24715731

  18. COS Side 2 NUV MAMA Fold Test

    NASA Astrophysics Data System (ADS)

    Bacinski, John

    2013-10-01

    The performance of the MAMA microchannel plate can be monitored using a MAMA fold analysis procedure. The fold analysis provides a measurement of the distribution of charge cloud sizes incident upon the anode giving some measure of changes in the pulse-height distribution of the MCP and, therefore, MCP gain. This proposal executes the same steps as the COS MAMA Fold Analysis {13128} during Cycle 20.This proposal is an exact duplication of nominal COS MAMA Fold Analysis {proposal 13128, Cycle 20}. Any changes 13128 or subsequent cycle submissions should be reflected in this proposal and vice versa.

  19. Protein folding using contact maps.

    PubMed

    Vendruscolo, M; Domany, E

    2000-01-01

    We discuss the problem of representations of protein structure and give the definition of contact maps. We present a method to obtain a three-dimensional polypeptide conformation from a contact map. We also explain how to deal with the case of nonphysical contact maps. We describe a stochastic method to perform dynamics in contact map space. We explain how the motion is restricted to physical regions of the space. First, we introduce the exact free energy of a contact map and discuss two simple approximations to it. Second, we present a method to derive energy parameters based on perception learning. We prove in an extensive number of situations that the pairwise contact approximation both when alone and when supplemented with a hydrophobic term is unsuitable for stabilizing proteins' native states. PMID:10668399

  20. Understanding Protein Folding from Advances of Fluorescence Energy Transfer.

    NASA Astrophysics Data System (ADS)

    Tcherkasskaya, Olga; Gronenborn, Angela M.

    2001-03-01

    A multi-site fluorescence energy transfer method was developed for the study of protein folding. Technique uses "tyrosine-phenylalanine" substitution mutagenesis to place the "tyrosine-guest" into positions of interest into the protein structure. Tetranitromethane modification of the tyrosine-guest renders this amino acid an acceptor of the tryptophan fluorescence. This approach can be applied to any protein system, and, most importantly, does not require single- or double-labeling of the protein molecule by a donor and/or an acceptor fluorophore. It is equally suited for equilibrium as well as kinetic studies of folding. We tested the methodology to monitor the equilibrium (un)folding of the immunoglobulin binding domain B1 of streptococcal protein G (GB1) induced by guanidine hydrochloride. Wild-type GB1 contains three tyrosines located at positions 3, 33, 45 and a single tryptophan residue at position 43. Two of the three tyrosines were replaced in turn, thereby allowing us to measure the energy transfer from Trp43 to each particular tyrosine. Overall, multi-parametrical experiments on GB1 including circular dichroism, steady state and time-resolved fluorescence, as well as fluorescence energy transfer revealed the existence of highly stable unfolded intermediates, which precede the formation of the rigid (native) secondary structure.

  1. WeFold: A Coopetition for Protein Structure Prediction

    PubMed Central

    Khoury, George A.; Liwo, Adam; Khatib, Firas; Zhou, Hongyi; Chopra, Gaurav; Bacardit, Jaume; Bortot, Leandro O.; Faccioli, Rodrigo A.; Deng, Xin; He, Yi; Krupa, Pawel; Li, Jilong; Mozolewska, Magdalena A.; Sieradzan, Adam K.; Smadbeck, James; Wirecki, Tomasz; Cooper, Seth; Flatten, Jeff; Xu, Kefan; Baker, David; Cheng, Jianlin; Delbem, Alexandre C. B.; Floudas, Christodoulos A.; Keasar, Chen; Levitt, Michael; Popović, Zoran; Scheraga, Harold A.; Skolnick, Jeffrey; Crivelli, Silvia N.; Players, Foldit

    2014-01-01

    The protein structure prediction problem continues to elude scientists. Despite the introduction of many methods, only modest gains were made over the last decade for certain classes of prediction targets. To address this challenge, a social-media based worldwide collaborative effort, named WeFold, was undertaken by thirteen labs. During the collaboration, the labs were simultaneously competing with each other. Here, we present the first attempt at “coopetition” in scientific research applied to the protein structure prediction and refinement problems. The coopetition was possible by allowing the participating labs to contribute different components of their protein structure prediction pipelines and create new hybrid pipelines that they tested during CASP10. This manuscript describes both successes and areas needing improvement as identified throughout the first WeFold experiment and discusses the efforts that are underway to advance this initiative. A footprint of all contributions and structures are publicly accessible at http://www.wefold.org. PMID:24677212

  2. Retinal and Choroidal Folds in Papilledema

    PubMed Central

    Sibony, Patrick A.; Kupersmith, Mark J.; Feldon, Steven E.; Wang, Jui-Kai; Garvin, Mona

    2015-01-01

    Purpose To determine the frequency, patterns, associations, and biomechanical implications of retinal and choroidal folds in papilledema due to idiopathic intracranial hypertension (IIH). Methods Retinal and choroidal folds were studied in patients enrolled in the IIH Treatment Trial using fundus photography (n = 165 study eyes) and spectral-domain optical coherence tomography (SD-OCT; n = 125). We examined the association between folds and peripapillary shape, retinal nerve fiber layer (RNFL) thickness, disc volume, Frisén grade, acuity, perimetric mean deviation, intraocular pressure, intracranial pressure, and refractive error. Results We identified three types of folds in IIH patients with papilledema: peripapillary wrinkles (PPW), retinal folds (RF), and choroidal folds (CF). Frequency, with photos, was 26%, 19%, and 1%, respectively; SD-OCT frequency was 46%, 47%, and 10%. At least one type of fold was present in 41% of patients with photos and 73% with SD-OCT. Spectral-domain OCT was more sensitive. Structural parameters related to the severity of papilledema were associated with PPW and RF, whereas anterior deformation of the peripapillary RPE/basement membrane layer was associated with CF and RF. Folds were not associated with vision loss at baseline. Conclusions Folds in papilledema are biomechanical signs of stress/strain on the optic nerve head and load-bearing structures induced by intracranial hypertension. Folds are best imaged with SD-OCT. The patterns of retinal and choroidal folds are the products of a complex interplay between the degree of papilledema and anterior deformation of the load-bearing structures (sclera and possibly the lamina cribrosa), both modulated by structural geometry and material properties of the optic nerve head. (ClinicalTrials.gov number, NCT01003639.) PMID:26335066

  3. [Choristoma in the vocal fold].

    PubMed

    Kempf, H-G; Störkel, S

    2005-04-01

    Choristoma in the larynx is rare and related to thyroid or glial tissue. The manifestation of salivary gland tissue in the larynx has not been reported to date. We present the case of an 80 year old male complaining of hoarseness and productive coughing. A left side tumourous swelling was seen in the larynx with intact vocal cord mobility. Using microlaryngoscopy the mass was resected without any intra- or postoperative problems. Histologically, a choristoma-heterotopic salivary gland tissue in the muscle and fat tissue--was found. The mucosa was intact and there were no signs of malignancy. Differential diagnosis of such masses in the larynx include benign lesions as well as specific infections, e.g. tuberculosis, sarcoidosis, amyloidosis and Wegener's granulomatosis. Especially in non-smokers, sarcoma, lymphoma and melanoma should be separated from the frequent squamous cell carcinoma of the laryngeal tissue. In rare cases, heterotopic tissue can mimic a tumourous mass. Intralaryngeal resection is the therapy of choice and should be recommended to the patient.

  4. Macromolecular Crowding Modulates Folding Mechanism of α/β Protein Apoflavodoxin

    NASA Astrophysics Data System (ADS)

    Homouz, D.; Stagg, L.; Wittungstafshede, P.; Cheung, M.

    2009-01-01

    Protein dynamics in cells may be different from that in dilute solutions in vitro since the environment in cells is highly concentrated with other macromolecules. This volume exclusion due to macromolecular crowding is predicted to affect both equilibrium and kinetic processes involving protein conformational changes. To quantify macromolecular crowding effects on protein folding mechanisms, here we have investigated the folding energy landscape of an alpha/beta protein, apoflavodoxin, in the presence of inert macromolecular crowding agents using in silico and in vitro approaches. By coarse-grained molecular simulations and topology-based potential interactions, we probed the effects of increased volume fraction of crowding agents (phi_c) as well as of crowding agent geometry (sphere or spherocylinder) at high phi_c. Parallel kinetic folding experiments with purified Desulfovibro desulfuricans apoflavodoxin in vitro were performed in the presence of Ficoll (sphere) and Dextran (spherocylinder) synthetic crowding agents. In conclusion, we have identified in silico crowding conditions that best enhance protein stability and discovered that upon manipulation of the crowding conditions, folding routes experiencing topological frustrations can be either enhanced or relieved. The test-tube experiments confirmed that apoflavodoxin's time-resolved folding path is modulated by crowding agent geometry. We propose that macromolecular crowding effects may be a tool for manipulation of protein folding and function in living cells.

  5. The molecular matching problem

    NASA Technical Reports Server (NTRS)

    Kincaid, Rex K.

    1993-01-01

    Molecular chemistry contains many difficult optimization problems that have begun to attract the attention of optimizers in the Operations Research community. Problems including protein folding, molecular conformation, molecular similarity, and molecular matching have been addressed. Minimum energy conformations for simple molecular structures such as water clusters, Lennard-Jones microclusters, and short polypeptides have dominated the literature to date. However, a variety of interesting problems exist and we focus here on a molecular structure matching (MSM) problem.

  6. Development and Application of a High Throughput Protein Unfolding Kinetic Assay

    PubMed Central

    Wang, Qiang; Waterhouse, Nicklas; Feyijinmi, Olusegun; Dominguez, Matthew J.; Martinez, Lisa M.; Sharp, Zoey; Service, Rachel; Bothe, Jameson R.; Stollar, Elliott J.

    2016-01-01

    The kinetics of folding and unfolding underlie protein stability and quantification of these rates provides important insights into the folding process. Here, we present a simple high throughput protein unfolding kinetic assay using a plate reader that is applicable to the studies of the majority of 2-state folding proteins. We validate the assay by measuring kinetic unfolding data for the SH3 (Src Homology 3) domain from Actin Binding Protein 1 (AbpSH3) and its stabilized mutants. The results of our approach are in excellent agreement with published values. We further combine our kinetic assay with a plate reader equilibrium assay, to obtain indirect estimates of folding rates and use these approaches to characterize an AbpSH3-peptide hybrid. Our high throughput protein unfolding kinetic assays allow accurate screening of libraries of mutants by providing both kinetic and equilibrium measurements and provide a means for in-depth ϕ-value analyses. PMID:26745729

  7. Local vs global motions in protein folding

    PubMed Central

    Maisuradze, Gia G.; Liwo, Adam; Senet, Patrick; Scheraga, Harold A.

    2013-01-01

    It is of interest to know whether local fluctuations in a polypeptide chain play any role in the mechanism by which the chain folds to the native structure of a protein. This question is addressed by analyzing folding and non-folding trajectories of a protein; as an example, the analysis is applied to the 37-residue triple β-strand WW domain from the Formin binding protein 28 (FBP28) (PDB ID: 1E0L). Molecular dynamics (MD) trajectories were generated with the coarse-grained united-residue force field, and one- and two-dimensional free-energy landscapes (FELs) along the backbone virtual-bond angle θ and backbone virtual-bond-dihedral angle γ of each residue, and principal components, respectively, were analyzed. The key residues involved in the folding of the FBP28 WW domain are elucidated by this analysis. The correlations between local and global motions are found. It is shown that most of the residues in the folding trajectories of the system studied here move in a concerted fashion, following the dynamics of the whole system. This demonstrates how the choice of a pathway has to involve concerted movements in order for this protein to fold. This finding also sheds light on the effectiveness of principal component analysis (PCA) for the description of the folding dynamics of the system studied. It is demonstrated that the FEL along the PCs, computed by considering only several critically-placed residues, can correctly describe the folding dynamics. PMID:23914144

  8. Protein folding: When ribosomes pick the structure

    NASA Astrophysics Data System (ADS)

    Sivertsson, Elin M.; Itzhaki, Laura S.

    2014-05-01

    Anfinsen's principle tells us that the folded structure of a protein is determined solely by its sequence. Now, it has been shown that the rate at which a polypeptide chain is synthesized in the cell can affect which of two alternative folded structures it adopts.

  9. Folding Polyominoes from One Level to Two

    ERIC Educational Resources Information Center

    Frederickson, Greg N.

    2011-01-01

    For any given polyomino, is it possible to cut it into pieces and then hinge the pieces, so that the polyomino folds up into a similar version of itself but two levels thick? While we don't know how to do this for every polyomino, the article does show how to cut, hinge, and fold polyominoes from several infinite classes, providing an…

  10. Using hydroxyl radical footprinting to explore the free energy landscape of protein folding

    PubMed Central

    Calabrese, Antonio N.; Ault, James R.; Radford, Sheena E.; Ashcroft, Alison E.

    2015-01-01

    Characterisation of the conformational states adopted during protein folding, including globally unfolded/disordered structures and partially folded intermediate species, is vital to gain fundamental insights into how a protein folds. In this work we employ fast photochemical oxidation of proteins (FPOP) to map the structural changes that occur in the folding of the four-helical bacterial immunity protein, Im7. Oxidative footprinting coupled with mass spectrometry (MS) is used to probe changes in the solvent accessibility of amino acid side-chains concurrent with the folding process, by quantifying the degree of oxidation experienced by the wild-type protein relative to a kinetically trapped, three-helical folding intermediate and an unfolded variant that lacks secondary structure. Analysis of the unfolded variant by FPOP–MS shows oxidative modifications consistent with the species adopting a solution conformation with a high degree of solvent accessibility. The folding intermediate, by contrast, experiences increased levels of oxidation relative to the wild-type, native protein only in regions destabilised by the amino acid substitutions introduced. The results demonstrate the utility of FPOP–MS to characterise protein variants in different conformational states and to provide insights into protein folding mechanisms that are complementary to measurements such as hydrogen/deuterium exchange labelling and Φ-value analysis. PMID:25746386

  11. Equilibrium Kinetic Network of the Villin Headpiece in Implicit Solvent

    PubMed Central

    Du, Weina; Bolhuis, Peter G.

    2015-01-01

    We applied the single-replica multiple-state transition-interface sampling method to elucidate the equilibrium kinetic network of the 35-residue-fragment (HP-35) villin headpiece in implicit water at room temperature. Starting from the native Protein Data Bank structure, nine (meta)stable states of the system were identified, from which the kinetic network was built by sampling pathways between these states. Application of transition path theory allowed analysis of the (un)folding mechanism. The resulting (un)folding rates agree well with experiments. This work demonstrates that high (un)folding barriers can now be studied. PMID:25606685

  12. Similarities between protein folding and granular jamming

    PubMed Central

    Jose, Prasanth P; Andricioaei, Ioan

    2012-01-01

    Grains and glasses, widely different materials, arrest their motions upon decreasing temperature and external load, respectively, in common ways, leading to a universal jamming phase diagram conjecture. However, unified theories are lacking, mainly because of the disparate nature of the particle interactions. Here we demonstrate that folded proteins exhibit signatures common to both glassiness and jamming by using temperature- and force-unfolding molecular dynamics simulations. Upon folding, proteins develop a peak in the interatomic force distributions that falls on a universal curve with experimentally measured forces on jammed grains and droplets. Dynamical signatures are found as a dramatic slowdown of stress relaxation upon folding. Together with granular similarities, folding is tied not just to the jamming transition, but a more nuanced picture of anisotropy, preparation protocol and internal interactions emerges. Results have implications for designing stable polymers and can open avenues to link protein folding to jamming theory. PMID:23093180

  13. Geometry and wetting of capillary folding.

    PubMed

    Péraud, Jean-Philippe; Lauga, Eric

    2014-04-01

    Capillary forces are involved in a variety of natural phenomena, ranging from droplet breakup to the physics of clouds. The forces from surface tension can also be exploited in industrial applications provided the length scales involved are small enough. Recent experimental investigations showed how to take advantage of capillarity to fold planar structures into three-dimensional configurations by selectively melting polymeric hinges joining otherwise rigid shapes. In this paper we use theoretical calculations to quantify the role of geometry and fluid wetting on the final folded state. Considering folding in two and three dimensions, studying both hydrophilic and hydrophobic situations with possible contact-angle hysteresis, and addressing the shapes to be folded to be successively infinite, finite, curved, kinked, and elastic, we are able to derive an overview of the geometrical parameter space available for capillary folding.

  14. Prediction of sound from human vocal folds

    NASA Astrophysics Data System (ADS)

    Bodony, Daniel; Luo, Haoxiang; Mittal, Rajat

    2007-11-01

    The creation of voiced sounds in humans depends on the flow-induced vibration of the vocal folds within the larynx. The vocal folds, which are a complex structural system of cartilage and tissue, create an oscillatory ``glottal jet'' whose harmonic content partially determines the tone of the speech. In this work we will discuss the process of sound generation in the laynx by combining a fully coupled two-dimensional fluid-structure simulation of the incompressible flow field in the vicinity of the vocal folds to an acoustic analogy description of the sound field. The structural dynamics of the vocal folds are based on physically realistic properties and are coupled to the motion of the fluid via an immersed boundary method. Relationships between the sound produced and the vocal fold dynamics will be discussed.

  15. Fan-fold shielded electrical leads

    DOEpatents

    Rohatgi, Rajeev R.; Cowan, Thomas E.

    1996-01-01

    Fan-folded electrical leads made from copper cladded Kapton, for example, with the copper cladding on one side serving as a ground plane and the copper cladding on the other side being etched to form the leads. The Kapton is fan folded with the leads located at the bottom of the fan-folds. Electrical connections are made by partially opening the folds of the fan and soldering, for example, the connections directly to the ground plane and/or the lead. The fan folded arrangement produces a number of advantages, such as electrically shielding the leads from the environment, is totally non-magnetic, and has a very low thermal conductivity, while being easy to fabricate.

  16. Fan-fold shielded electrical leads

    DOEpatents

    Rohatgi, R.R.; Cowan, T.E.

    1996-06-11

    Disclosed are fan-folded electrical leads made from copper cladded Kapton, for example, with the copper cladding on one side serving as a ground plane and the copper cladding on the other side being etched to form the leads. The Kapton is fan folded with the leads located at the bottom of the fan-folds. Electrical connections are made by partially opening the folds of the fan and soldering, for example, the connections directly to the ground plane and/or the lead. The fan folded arrangement produces a number of advantages, such as electrically shielding the leads from the environment, is totally non-magnetic, and has a very low thermal conductivity, while being easy to fabricate. 3 figs.

  17. Folding and Finding RNA Secondary Structure

    PubMed Central

    Mathews, David H.; Moss, Walter N.; Turner, Douglas H.

    2010-01-01

    SUMMARY Optimal exploitation of the expanding database of sequences requires rapid finding and folding of RNAs. Methods are reviewed that automate folding and discovery of RNAs with algorithms that couple thermodynamics with chemical mapping, NMR, and/or sequence comparison. New functional noncoding RNAs in genome sequences can be found by combining sequence comparison with the assumption that functional noncoding RNAs will have more favorable folding free energies than other RNAs. When a new RNA is discovered, experiments and sequence comparison can restrict folding space so that secondary structure can be rapidly determined with the help of predicted free energies. In turn, secondary structure restricts folding in three dimensions, which allows modeling of three-dimensional structure. An example from a domain of a retrotransposon is described. Discovery of new RNAs and their structures will provide insights into evolution, biology, and design of therapeutics. Applications to studies of evolution are also reviewed. PMID:20685845

  18. The robustness and innovability of protein folds.

    PubMed

    Tóth-Petróczy, Agnes; Tawfik, Dan S

    2014-06-01

    Assignment of protein folds to functions indicates that >60% of folds carry out one or two enzymatic functions, while few folds, for example, the TIM-barrel and Rossmann folds, exhibit hundreds. Are there structural features that make a fold amenable to functional innovation (innovability)? Do these features relate to robustness--the ability to readily accumulate sequence changes? We discuss several hypotheses regarding the relationship between the architecture of a protein and its evolutionary potential. We describe how, in a seemingly paradoxical manner, opposite properties, such as high stability and rigidity versus conformational plasticity and structural order versus disorder, promote robustness and/or innovability. We hypothesize that polarity--differentiation and low connectivity between a protein's scaffold and its active-site--is a key prerequisite for innovability.

  19. Zinc Binding Modulates the Entire Folding Free Energy Surface of Human Cu,Zn Superoxide Dismutase

    PubMed Central

    Kayatekin, Can; Zitzewitz, Jill A.; Matthews, C. Robert

    2009-01-01

    Over 100 amino acid replacements in human Cu, Zn superoxide dismutase (SOD) are known to cause amyotrophic lateral sclerosis, a gain-of-function neurodegenerative disease that destroys motor neurons. Supposing that aggregates of partially-folded states are primarily responsible for toxicity, the role of the structurally-important zinc ion in defining the folding free energy surface of dimeric SOD was determined by comparing the thermodynamic and kinetic folding properties of the zinc-free and zinc-bound forms of the protein. The presence of zinc was found to decrease the free energies of a peptide model of the unfolded monomer, a stable variant of the folded monomeric intermediate and the folded dimeric species. The unfolded state binds zinc weakly with a micromolar dissociation constant, and the folded monomeric intermediate and the native dimeric form both bind zinc tightly, with sub-nanomolar dissociation constants. Coupled with the strong driving force for the subunit association reaction, the shift in the populations towards more well-folded states in the presence of zinc decreases the steady-state populations of higher-energy states in SOD under expected in vivo zinc concentrations (∼nanomolar). The significant decrease in the population of partially-folded states is expected to diminish their potential for aggregation and account for the known protective effect of zinc. The ∼100-fold increase in the rate of folding of SOD in the presence of micromolar concentrations of zinc demonstrates a significant role for a pre-organized zinc-binding loop in the transition state ensemble for the rate-limiting monomer folding reaction in this β-barrel protein. PMID:18840448

  20. Some current problems in oxidation kinetics

    NASA Technical Reports Server (NTRS)

    Benson, S. W.

    1972-01-01

    Experimental data in low temperature and high temperature oxidations are examined from the point of view of reported quantitative inconsistencies. Activation energies for tBuO2 metathesis reactions with alkanes appear to be 7 kcal/mole higher than for comparable reactions of HO2. Related isomerization reactions are examined in the light of these differences without reaching any simple conclusions. The Russell mechanism for a 6-membered, cyclic, transition state for termination of primary and secondary alkyl peroxy radicals is shown to be either inconsistent with thermochemical data, or else unique to solution reactions. Addition reactions of O3 with olefins and acetylenes are shown thermochemically to have the possibility of following concerted and biradical pathways, respectively. Recent data showing strong inhibition by PbO coated surfaces of both oxidation and pyrolysis of i-C4H10 are examined in terms of mechanism.

  1. Multiple-probe analysis of folding and unfolding pathways of human serum albumin. Evidence for a framework mechanism of folding.

    PubMed

    Santra, Manas Kumar; Banerjee, Abhijit; Krishnakumar, Shyam Sundar; Rahaman, Obaidur; Panda, Dulal

    2004-05-01

    The changes in the far-UV CD signal, intrinsic tryptophan fluorescence and bilirubin absorbance showed that the guanidine hydrochloride (GdnHCl)-induced unfolding of a multidomain protein, human serum albumin (HSA), followed a two-state process. However, using environment sensitive Nile red fluorescence, the unfolding and folding pathways of HSA were found to follow a three-state process and an intermediate was detected in the range 0.25-1.5 m GdnHCl. The intermediate state displayed 45% higher fluorescence intensity than that of the native state. The increase in the Nile red fluorescence was found to be due to an increase in the quantum yield of the HSA-bound Nile red. Low concentrations of GdnHCl neither altered the binding affinity of Nile red to HSA nor induced the aggregation of HSA. In addition, the secondary structure of HSA was not perturbed during the first unfolding transition (<1.5 m GdnHCl); however, the secondary structure was completely lost during the second transition. The data together showed that the half maximal loss of the tertiary structure occurred at a lower GdnHCl concentration than the loss of the secondary structure. Further kinetic studies of the refolding process of HSA using multiple spectroscopic techniques showed that the folding occurred in two phases, a burst phase followed by a slow phase. An intermediate with native-like secondary structure but only a partial tertiary structure was found to form in the burst phase of refolding. Then, the intermediate slowly folded into the native state. An analysis of the refolding data suggested that the folding of HSA could be best explained by the framework model.

  2. Dynamics of an Ultrafast Folding Subdomain in the Context of a Larger Protein Fold

    PubMed Central

    Davis, Caitlin M.; Dyer, R. Brian

    2014-01-01

    Small fast folding subdomains with low contact order have been postulated to facilitate the folding of larger proteins. We have tested this idea by determining how the fastest folding linear β-hairpin, CLN025, which folds on the nanosecond time scale, folds within the context of a two-hairpin WW domain system, which folds on the microsecond time scale. The folding of the wild type FBP28 WW domain was compared to constructs in which each of the loops was replaced by CLN025. A combination of FTIR spectroscopy and laser-induced temperature-jump coupled with infrared spectroscopy was used to probe changes in the peptide backbone. The relaxation dynamics of the β-sheets and β-turn were measured independently by probing the corresponding bands assigned in the amide I region. The folding rate of the CLN025 β-hairpin is unchanged within the larger protein. Insertion of the β-hairpin into the second loop results in an overall stabilization of the WW domain and a relaxation lifetime five times faster than the parent WW domain. In both mutants, folding is initiated in the turns and the β-sheets form last. These results demonstrate that fast folding subdomains can be used to speed the folding of more complex proteins, and that the folding dynamics of the subdomain is unchanged within the context of the larger protein. PMID:24320936

  3. Effects of Folding on Metalloprotein Active Sites

    NASA Astrophysics Data System (ADS)

    Winkler, Jay R.; Wittung-Stafshede, Pernilla; Leckner, Johan; Malmstrom, Bo G.; Gray, Harry B.

    1997-04-01

    Experimental data for the unfolding of cytochrome c and azurin by guanidinium chloride (GuHCl) are used to construct free-energy diagrams for the folding of the oxidized and reduced proteins. With cytochrome c, the driving force for folding the reduced protein is larger than that for the oxidized form. Both the oxidized and the reduced folded forms of yeast cytochrome c are less stable than the corresponding states of the horse protein. Due to the covalent attachment of the heme and its fixed tetragonal coordination geometry, cytochrome c folding can be described by a two-state model. A thermodynamic cycle leads to an expression for the difference in self-exchange reorganization energies for the folded and unfolded proteins. The reorganization energy for electron exchange in the folded protein is approximately 0.5 eV smaller than that for a heme in aqueous solution. The finding that reduced azurin unfolds at lower GuHCl concentrations than the oxidized protein suggests that the coordination structure of copper is different in oxidized and reduced unfolded states: it is likely that the geometry of CuI in the unfolded protein is linear or trigonal, whereas CuII prefers to be tetragonal. The evidence indicates that protein folding lowers the azurin reorganization energy by roughly 1.7 eV relative to an aqueous Cu(1,10-phenanthroline)2{}2+/+ reference system.

  4. Quantification of a Helical Origami Fold

    NASA Astrophysics Data System (ADS)

    Dai, Eric; Han, Xiaomin; Chen, Zi

    2015-03-01

    Origami, the Japanese art of paper folding, is traditionally viewed as an amusing pastime and medium of artistic expression. However, in recent years, origami has served as a source of inspiration for innovations in science and engineering. Here, we present the geometric and mechanical properties of a twisting origami fold. The origami structure created by the fold exhibits several interesting properties, including rigid foldibility, local bistability and finely tunable helical coiling, with control over pitch, radius and handedness of the helix. In addition, the pattern generated by the fold closely mimics the twist buckling patterns shown by thin materials, for example, a mobius strip. We use six parameters of the twisting origami pattern to generate a fully tunable graphical model of the fold. Finally, we present a mathematical model of the local bistability of the twisting origami fold. Our study elucidates the mechanisms behind the helical coiling and local bistability of the twisting origami fold, with potential applications in robotics and deployable structures. Acknowledgment to Branco Weiss Fellowship for funding.

  5. Predicting protein folding rate change upon point mutation using residue-level coevolutionary information.

    PubMed

    Mallik, Saurav; Das, Smita; Kundu, Sudip

    2016-01-01

    Change in folding kinetics of globular proteins upon point mutation is crucial to a wide spectrum of biological research, such as protein misfolding, toxicity, and aggregations. Here we seek to address whether residue-level coevolutionary information of globular proteins can be informative to folding rate changes upon point mutations. Generating residue-level coevolutionary networks of globular proteins, we analyze three parameters: relative coevolution order (rCEO), network density (ND), and characteristic path length (CPL). A point mutation is considered to be equivalent to a node deletion of this network and respective percentage changes in rCEO, ND, CPL are found linearly correlated (0.84, 0.73, and -0.61, respectively) with experimental folding rate changes. The three parameters predict the folding rate change upon a point mutation with 0.031, 0.045, and 0.059 standard errors, respectively.

  6. Folding simulations of alanine-based peptides with lysine residues.

    PubMed Central

    Sung, S S

    1995-01-01

    The folding of short alanine-based peptides with different numbers of lysine residues is simulated at constant temperature (274 K) using the rigid-element Monte Carlo method. The solvent-referenced potential has prevented the multiple-minima problem in helix folding. From various initial structures, the peptides with three lysine residues fold into helix-dominated conformations with the calculated average helicity in the range of 60-80%. The peptide with six lysine residues shows only 8-14% helicity. These results agree well with experimental observations. The intramolecular electrostatic interaction of the charged lysine side chains and their electrostatic hydration destabilize the helical conformations of the peptide with six lysine residues, whereas these effects on the peptides with three lysine residues are small. The simulations provide insight into the helix-folding mechanism, including the beta-bend intermediate in helix initiation, the (i, i + 3) hydrogen bonds, the asymmetrical helix propagation, and the asymmetrical helicities in the N- and C-terminal regions. These findings are consistent with previous studies. PMID:7756550

  7. Local analysis near a folded saddle-node singularity

    NASA Astrophysics Data System (ADS)

    Krupa, Martin; Wechselberger, Martin

    Folded saddle-nodes occur generically in one parameter families of singularly perturbed systems with two slow variables. We show that these folded singularities are the organizing centers for two main delay phenomena in singular perturbation problems: canards and delayed Hopf bifurcations. We combine techniques from geometric singular perturbation theory—the blow-up technique—and from delayed Hopf bifurcation theory—complex time path analysis—to analyze the flow near such folded saddle-nodes. In particular, we show the existence of canards as intersections of stable and unstable slow manifolds. To derive these canard results, we extend the singularly perturbed vector field into the complex domain and study it along elliptic paths. This enables us to extend the invariant slow manifolds beyond points where normal hyperbolicity is lost. Furthermore, we define a way-in/way-out function describing the maximal delay expected for generic solutions passing through a folded saddle-node singularity. Branch points associated with the change from a complex to a real eigenvalue structure in the variational equation along the critical (slow) manifold make our analysis significantly different from the classical delayed Hopf bifurcation analysis where these eigenvalues are complex only.

  8. Osteochondrodysplasia in three Scottish Fold cats.

    PubMed

    Chang, Jinhwa; Jung, Joohyun; Oh, Sunkyoung; Lee, Sungok; Kim, Gyeongmin; Kim, Haksang; Kweon, Ohkyeong; Yoon, Junghee; Choi, Mincheol

    2007-09-01

    This report explains typical radiographic features of Scottish Fold osteochondrodysplasia. Three Scottish Fold cats suffering from lameness were referred to the Veterinary Medical Teaching Hospital, Seoul National University, Korea. Based on the breed predisposition, history, clinical signs, physical examination, and radiographic findings, Scottish Fold osteochondrodysplasia was confirmed in three cases. Radiographic changes mainly included exostosis and secondary arthritis around affected joint lesions, and defective conformation in the phalanges and caudal vertebrae. The oral chondroprotective agents such as glucosamine and chondroitin sulfate make the patients alleviate their pain without adverse effects. PMID:17679781

  9. Mechanical Models of Fault-Related Folding

    SciTech Connect

    Johnson, A. M.

    2003-01-09

    The subject of the proposed research is fault-related folding and ground deformation. The results are relevant to oil-producing structures throughout the world, to understanding of damage that has been observed along and near earthquake ruptures, and to earthquake-producing structures in California and other tectonically-active areas. The objectives of the proposed research were to provide both a unified, mechanical infrastructure for studies of fault-related foldings and to present the results in computer programs that have graphical users interfaces (GUIs) so that structural geologists and geophysicists can model a wide variety of fault-related folds (FaRFs).

  10. Kinetic buffers.

    PubMed

    Alibrandi, Giuseppe; Fabbrizzi, Luigi; Licchelli, Maurizio; Puglisi, Antonio

    2015-01-12

    This paper proposes a new type of molecular device that is able to act as an inverse proton sponge to slowly decrease the pH inside a reaction vessel. This makes the automatic monitoring of the concentration of pH-sensitive systems possible. The device is a composite formed of an alkyl chloride, which kinetically produces acidity, and a buffer that thermodynamically modulates the variation in pH value. Profiles of pH versus time (pH-t plots) have been generated under various experimental conditions by computer simulation, and the device has been tested by carrying out automatic spectrophotometric titrations, without using an autoburette. To underline the wide variety of possible applications, this new system has been used to realize and monitor HCl uptake by a di-copper(II) bistren complex in a single run, in a completely automatic experiment.

  11. Distinct Contribution of Electrostatics, Initial Conformational Ensemble, and Macromolecular Stability in RNA Folding

    SciTech Connect

    Laederach,A.; Shcherbakova, I.; Jonikas, M.; Altman, R.; Brenowitz, M.

    2007-01-01

    We distinguish the contribution of the electrostatic environment, initial conformational ensemble, and macromolecular stability on the folding mechanism of a large RNA using a combination of time-resolved 'Fast Fenton' hydroxyl radical footprinting and exhaustive kinetic modeling. This integrated approach allows us to define the folding landscape of the L-21 Tetrahymena thermophila group I intron structurally and kinetically from its earliest steps with unprecedented accuracy. Distinct parallel pathways leading the RNA to its native form upon its Mg2+-induced folding are observed. The structures of the intermediates populating the pathways are not affected by variation of the concentration and type of background monovalent ions (electrostatic environment) but are altered by a mutation that destabilizes one domain of the ribozyme. Experiments starting from different conformational ensembles but folding under identical conditions show that whereas the electrostatic environment modulates molecular flux through different pathways, the initial conformational ensemble determines the partitioning of the flux. This study showcases a robust approach for the development of kinetic models from collections of local structural probes.

  12. Under-folded proteins: Conformational ensembles and their roles in protein folding, function, and pathogenesis.

    PubMed

    Uversky, Vladimir N

    2013-11-01

    For decades, protein function was intimately linked to the presence of a unique, aperiodic crystal-like structure in a functional protein. The two only places for conformational ensembles of under-folded (or partially folded) protein forms in this picture were either the end points of the protein denaturation processes or transiently populated folding intermediates. Recent years witnessed dramatic change in this perception and conformational ensembles, which the under-folded proteins are, have moved from the shadow. Accumulated to date data suggest that a protein can exist in at least three global forms-functional and folded, functional and intrinsically disordered (nonfolded), and nonfunctional and misfolded/aggregated. Under-folded protein states are crucial for each of these forms, serving as important folding intermediates of ordered proteins, or as functional states of intrinsically disordered proteins (IDPs) and IDP regions (IDPRs), or as pathology triggers of misfolded proteins. Based on these observations, conformational ensembles of under-folded proteins can be classified as transient (folding and misfolding intermediates) and permanent (IDPs and stable misfolded proteins). Permanently under-folded proteins can further be split into intentionally designed (IDPs and IDPRs) and unintentionally designed (misfolded proteins). Although intrinsic flexibility, dynamics, and pliability are crucial for all under-folded proteins, the different categories of under-foldedness are differently encoded in protein amino acid sequences.

  13. Dew-driven folding of insect wings

    NASA Astrophysics Data System (ADS)

    Dickerson, Andrew; Beadles, Sam; Clement, Courtney; Hu, David

    2013-11-01

    Small insect wings fold into tacos when exposed to dewfall or fog for extended times. Such shapes are tightly held together and require great force or long evaporation times for the wings to unfold. In this experimental investigation, we use time-lapse and high-speed videography on a mosquito wing exposed to fog to characterize the folding process from a flat wing to a taco. We observe a taco is formed through a series of processes involving wing bending, unbending, and subsequent tight folding of the wing following the sliding of the drop off the wing. We use a simplified 2D model to determine the forces coalescing drops exert on the wing, and present folding-resistant design suggestions for micro-aerial vehicle wings.

  14. Folded waveguide coupler for ion cyclotron heating

    SciTech Connect

    Owens, T.L.; Chen, G.L.

    1986-01-01

    A new type of waveguide coupler for plasma heating in the ion cyclotron range of frequencies is described. The coupler consists of a series of interleaved metallic vanes within a rectangular enclosure analogous to a wide rectangular waveguide that has been ''folded'' several times. At the mouth of the coupler, a plate is attached which contains coupling apertures in each fold or every other fold of the waveguide, depending upon the wavenumber spectrum desired. This plate serves primarily as a wave field polarizer that converts coupler fields to the polarization of the fast magnetosonic wave within the plasma. Theoretical estimates indicate that the folded waveguide is capable of high-efficiency, multimegawatt operation into a plasma. Bench tests have verified the predicted field structure within the waveguide in preparation for high-power tests on the Radio Frequency Test Facility at the Oak Ridge National Laboratory.

  15. Self-folding miniature elastic electric devices

    NASA Astrophysics Data System (ADS)

    Miyashita, Shuhei; Meeker, Laura; Tolley, Michael T.; Wood, Robert J.; Rus, Daniela

    2014-09-01

    Printing functional materials represents a considerable impact on the access to manufacturing technology. In this paper we present a methodology and validation of print-and-self-fold miniature electric devices. Polyvinyl chloride laminated sheets based on metalized polyester film show reliable self-folding processes under a heat application, and it configures 3D electric devices. We exemplify this technique by fabricating fundamental electric devices, namely a resistor, capacitor, and inductor. Namely, we show the development of a self-folded stretchable resistor, variable resistor, capacitive strain sensor, and an actuation mechanism consisting of a folded contractible solenoid coil. Because of their pre-defined kinematic design, these devices feature elasticity, making them suitable as sensors and actuators in flexible circuits. Finally, an RLC circuit obtained from the integration of developed devices is demonstrated, in which the coil based actuator is controlled by reading a capacitive strain sensor.

  16. Reinke Edema: Watch For Vocal Fold Cysts.

    PubMed

    Tüzüner, Arzu; Demirci, Sule; Yavanoglu, Ahmet; Kurkcuoglu, Melih; Arslan, Necmi

    2015-06-01

    Reinke edema is one of the common cause of dysphonia middle-aged population, and severe thickening of vocal folds require surgical treatment. Smoking plays a major role on etiology. Vocal fold cysts are also benign lesions and vocal trauma blamed for acquired cysts. We would like to present 3 cases with vocal fold cyst related with Reinke edema. First case had a subepidermal epidermoid cyst with Reinke edema, which could be easily observed before surgery during laryngostroboscopy. Second case had a mucous retention cyst into the edematous Reinke tissue, which was detected during surgical intervention, and third case had a epidermoid cyst that occurred 2 months after before microlaryngeal operation regarding Reinke edema reduction. These 3 cases revealed that surgical management of Reinke edema needs a careful dissection and close follow-up after surgery for presence of vocal fold cysts.

  17. Statistical properties of a folded elastic rod

    NASA Astrophysics Data System (ADS)

    Bayart, Elsa; Deboeuf, Stéphanie; Boué, Laurent; Corson, Francis; Boudaoud, Arezki; Adda-Bedia, Mokhtar

    2010-03-01

    A large variety of elastic structures naturally seem to be confined into environments too small to accommodate them; the geometry of folded structures span a wide range of length-scales. The elastic properties of these confined systems are further constrained by self-avoidance as well as by the dimensionality of both structures and container. To mimic crumpled paper, we devised an experimental setup to study the packing of a dimensional elastic object in 2D geometries: an elastic rod is folded at the center of a circular Hele-Shaw cell by a centripetal force. The initial configuration of the rod and the acceleration of the rotating disk allow to span different final folded configurations while the final rotation speed controls the packing intensity. Using image analysis we measure geometrical and mechanical properties of the folded configurations, focusing on length, curvature and energy distributions.

  18. Monster Mash: Protein Folding Gone Wrong

    MedlinePlus

    ... Articles | Inside Life Science Home Page Monster Mash: Protein Folding Gone Wrong By Joseph Piergrossi Posted October 31, 2013 In this image, globs of misfolded proteins called amyloid plaques (blobs) are found outside neurons ( ...

  19. Unexplained Profound Hypoglycemia After Vocal Fold Lipoinjection.

    PubMed

    Modanlou, Shohreh; Marie Giglio, Nicole; Carroll, Thomas; Pancaro, Carlo

    2016-02-01

    Vocal fold injection is used for the management of glottal incompetence from various causes. The procedure is well tolerated and has few reported complications. We present a case of a 66-year-old man with long-lasting hoarseness secondary to vocal fold atrophy, who underwent an uneventful bilateral vocal fold injection with autologous fat. While in the recovery area, he experienced profuse sweating approximately 30 minutes after the surgical procedure. His blood glucose value was measured at 24 mg/dL, and plasmatic insulin level was 246 mU/L. To our knowledge, this is the first reported case of a systemic side effect after vocal fold lipoinjection. PMID:26491839

  20. Topology Explains Why Automobile Sunshades Fold Oddly

    ERIC Educational Resources Information Center

    Feist, Curtis; Naimi, Ramin

    2009-01-01

    Automobile sunshades always fold into an "odd" number of loops. The explanation why involves elementary topology (braid theory and linking number, both explained in detail here with definitions and examples), and an elementary fact from algebra about symmetric group.

  1. The hydrogen exchange core and protein folding.

    PubMed Central

    Li, R.; Woodward, C.

    1999-01-01

    A database of hydrogen-deuterium exchange results has been compiled for proteins for which there are published rates of out-exchange in the native state, protection against exchange during folding, and out-exchange in partially folded forms. The question of whether the slow exchange core is the folding core (Woodward C, 1993, Trends Biochem Sci 18:359-360) is reexamined in a detailed comparison of the specific amide protons (NHs) and the elements of secondary structure on which they are located. For each pulsed exchange or competition experiment, probe NHs are shown explicitly; the large number and broad distribution of probe NHs support the validity of comparing out-exchange with pulsed-exchange/competition experiments. There is a strong tendency for the same elements of secondary structure to carry NHs most protected in the native state, NHs first protected during folding, and NHs most protected in partially folded species. There is not a one-to-one correspondence of individual NHs. Proteins for which there are published data for native state out-exchange and theta values are also reviewed. The elements of secondary structure containing the slowest exchanging NHs in native proteins tend to contain side chains with high theta values or be connected to a turn/loop with high theta values. A definition for a protein core is proposed, and the implications for protein folding are discussed. Apparently, during folding and in the native state, nonlocal interactions between core sequences are favored more than other possible nonlocal interactions. Other studies of partially folded bovine pancreatic trypsin inhibitor (Barbar E, Barany G, Woodward C, 1995, Biochemistry 34:11423-11434; Barber E, Hare M, Daragan V, Barany G, Woodward C, 1998, Biochemistry 37:7822-7833), suggest that developing cores have site-specific energy barriers between microstates, one disordered, and the other(s) more ordered. PMID:10452602

  2. [Surgery of benign vocal fold lesions].

    PubMed

    Olthoff, A

    2016-09-01

    Surgical treatment of benign vocal fold lesions can be indicated for clinical or functional reasons. The principles of phonosurgery have to be maintained in either case. The appropriate phonosurgical technique depends on the type of vocal fold lesion. Depending on the findings, phonosurgery aims to maintain or improve voice quality. The evaluation of clinical and functional results includes indirect laryngoscopy, videostroboscopy, and voice analysis. PMID:27552826

  3. On a smooth quintic 4-fold

    SciTech Connect

    Cheltsov, I A

    2000-10-31

    The birational geometry of an arbitrary smooth quintic 4-fold is studied using the properties of log pairs. As a result, a new proof of its birational rigidity is given and all birational maps of a smooth quintic 4-fold into fibrations with general fibre of Kodaira dimension zero are described. In the Addendum similar results are obtained for all smooth hypersurfaces of degree n in P{sup n} in the case of n equal to 6, 7, or 8.

  4. Molecular crowders and cosolutes promote folding cooperativity of RNA under physiological ionic conditions.

    PubMed

    Strulson, Christopher A; Boyer, Joshua A; Whitman, Elisabeth E; Bevilacqua, Philip C

    2014-03-01

    Folding mechanisms of functional RNAs under idealized in vitro conditions of dilute solution and high ionic strength have been well studied. Comparatively little is known, however, about mechanisms for folding of RNA in vivo where Mg(2+) ion concentrations are low, K(+) concentrations are modest, and concentrations of macromolecular crowders and low-molecular-weight cosolutes are high. Herein, we apply a combination of biophysical and structure mapping techniques to tRNA to elucidate thermodynamic and functional principles that govern RNA folding under in vivo-like conditions. We show by thermal denaturation and SHAPE studies that tRNA folding cooperativity increases in physiologically low concentrations of Mg(2+) (0.5-2 mM) and K(+) (140 mM) if the solution is supplemented with physiological amounts (∼ 20%) of a water-soluble neutral macromolecular crowding agent such as PEG or dextran. Low-molecular-weight cosolutes show varying effects on tRNA folding cooperativity, increasing or decreasing it based on the identity of the cosolute. For those additives that increase folding cooperativity, the gain is manifested in sharpened two-state-like folding transitions for full-length tRNA over its secondary structural elements. Temperature-dependent SHAPE experiments in the absence and presence of crowders and cosolutes reveal extent of cooperative folding of tRNA on a nucleotide basis and are consistent with the melting studies. Mechanistically, crowding agents appear to promote cooperativity by stabilizing tertiary structure, while those low molecular cosolutes that promote cooperativity stabilize tertiary structure and/or destabilize secondary structure. Cooperative folding of functional RNA under physiological-like conditions parallels the behavior of many proteins and has implications for cellular RNA folding kinetics and evolution.

  5. Folding dynamics of Trp-cage in the presence of chemical interference and macromolecular crowding. I

    NASA Astrophysics Data System (ADS)

    Samiotakis, Antonios; Cheung, Margaret S.

    2011-11-01

    Proteins fold and function in the crowded environment of the cell's interior. In the recent years it has been well established that the so-called "macromolecular crowding" effect enhances the folding stability of proteins by destabilizing their unfolded states for selected proteins. On the other hand, chemical and thermal denaturation is often used in experiments as a tool to destabilize a protein by populating the unfolded states when probing its folding landscape and thermodynamic properties. However, little is known about the complicated effects of these synergistic perturbations acting on the kinetic properties of proteins, particularly when large structural fluctuations, such as protein folding, have been involved. In this study, we have first investigated the folding mechanism of Trp-cage dependent on urea concentration by coarse-grained molecular simulations where the impact of urea is implemented into an energy function of the side chain and/or backbone interactions derived from the all-atomistic molecular dynamics simulations with urea through a Boltzmann inversion method. In urea solution, the folding rates of a model miniprotein Trp-cage decrease and the folded state slightly swells due to a lack of contact formation between side chains at the terminal regions. In addition, the equilibrium m-values of Trp-cage from the computer simulations are in agreement with experimental measurements. We have further investigated the combined effects of urea denaturation and macromolecular crowding on Trp-cage's folding mechanism where crowding agents are modeled as hard-spheres. The enhancement of folding rates of Trp-cage is most pronounced by macromolecular crowding effect when the extended conformations of Trp-cast dominate at high urea concentration. Our study makes quantitatively testable predictions on protein folding dynamics in a complex environment involving both chemical denaturation and macromolecular crowding effects.

  6. Muscular Anatomy of the Human Ventricular Folds

    PubMed Central

    Moon, Jerald; Alipour, Fariborz

    2013-01-01

    Objective The purpose of this study was to better understand the muscular anatomy of the ventricular folds (VF) to help improve biomechanical modeling of phonation and to better understand the role of these muscles during phonatory and non-phonatory tasks. Method Four human larynges were decalcified and sectioned coronally from the posterior to anterior using a CryoJane tape transfer system, and stained using Massons trichrome. The total and relative area of muscles observed in each section were calculated and used for characterizing muscle distribution within the ventricular folds. Results The ventricular folds of the larynges contained anteriorly coursing thyroarytenoid and ventricularis muscle fibers lying in the lower half of the VF posteriorly, with some ventricularis muscle evident in the upper and lateral portion of the fold more anteriorly. Very little muscle tissue was observed in the medial half of the fold, and the anterior half of the VF was largely devoid of any muscle tissue. All four VF’s contained muscle bundles coursing superiorly and medially through the upper half of the fold toward the lateral margin of the epiglottis. Conclusions While variability in expression was evident, the well-defined thyroarytenoid muscle was readily apparent lateral to the arytenoid cartilage in all specimens. PMID:24224399

  7. Protein Folding and Misfolding on Surfaces

    PubMed Central

    Stefani, Massimo

    2008-01-01

    Protein folding, misfolding and aggregation, as well as the way misfolded and aggregated proteins affects cell viability are emerging as key themes in molecular and structural biology and in molecular medicine. Recent advances in the knowledge of the biophysical basis of protein folding have led to propose the energy landscape theory which provides a consistent framework to better understand how a protein folds rapidly and efficiently to the compact, biologically active structure. The increased knowledge on protein folding has highlighted its strict relation to protein misfolding and aggregation, either process being in close competition with the other, both relying on the same physicochemical basis. The theory has also provided information to better understand the structural and environmental factors affecting protein folding resulting in protein misfolding and aggregation into ordered or disordered polymeric assemblies. Among these, particular importance is given to the effects of surfaces. The latter, in some cases make possible rapid and efficient protein folding but most often recruit proteins/peptides increasing their local concentration thus favouring misfolding and accelerating the rate of nucleation. It is also emerging that surfaces can modify the path of protein misfolding and aggregation generating oligomers and polymers structurally different from those arising in the bulk solution and endowed with different physical properties and cytotoxicities. PMID:19330090

  8. Structural Characteristics of Novel Protein Folds

    PubMed Central

    Fernandez-Fuentes, Narcis; Dybas, Joseph M.; Fiser, Andras

    2010-01-01

    Folds are the basic building blocks of protein structures. Understanding the emergence of novel protein folds is an important step towards understanding the rules governing the evolution of protein structure and function and for developing tools for protein structure modeling and design. We explored the frequency of occurrences of an exhaustively classified library of supersecondary structural elements (Smotifs), in protein structures, in order to identify features that would define a fold as novel compared to previously known structures. We found that a surprisingly small set of Smotifs is sufficient to describe all known folds. Furthermore, novel folds do not require novel Smotifs, but rather are a new combination of existing ones. Novel folds can be typified by the inclusion of a relatively higher number of rarely occurring Smotifs in their structures and, to a lesser extent, by a novel topological combination of commonly occurring Smotifs. When investigating the structural features of Smotifs, we found that the top 10% of most frequent ones have a higher fraction of internal contacts, while some of the most rare motifs are larger, and contain a longer loop region. PMID:20421995

  9. Optimum folding pathways for growing protein chains.

    PubMed

    Senturk, Serife; Baday, Sefer; Arkun, Yaman; Erman, Burak

    2007-11-26

    The folding of a protein is studied as it grows residue by residue from the N-terminus and enters an environment that stabilizes the folded state. This mode of folding of a growing chain is different from refolding where the full chain folds from a disordered initial configuration to the native state. We propose a sequential dynamic optimization method that computes the evolution of optimum folding pathways as amino acid residues are added to the peptide chain one by one. The dynamic optimization formulation is deterministic and uses Newton's equations of motion and a Go-type potential that establishes the native contacts and excluded volume effects. The method predicts the optimal energy-minimizing path among all the alternative feasible pathways. As two examples, the folding of the chicken villin headpiece, a 36-residue protein, and chymotrypsin inhibitor 2 (CI2), a 64-residue protein, are studied. Results on the villin headpiece show significant differences from the refolding of the same chain studied previously. Results on CI2 mostly agree with the results of refolding experiments and computational work.

  10. "Wet" Versus "Dry" Folding of Polyproline

    NASA Astrophysics Data System (ADS)

    Shi, Liuqing; Holliday, Alison E.; Bohrer, Brian C.; Kim, Doyong; Servage, Kelly A.; Russell, David H.; Clemmer, David E.

    2016-06-01

    When the all- cis polyproline-I helix (PPI, favored in 1-propanol) of polyproline-13 is introduced into water, it folds into the all- trans polyproline-II (PPII) helix through at least six intermediates [Shi, L., Holliday, A.E., Shi, H., Zhu, F., Ewing, M.A., Russell, D.H., Clemmer, D.E.: Characterizing intermediates along the transition from PPI to PPII using ion mobility-mass spectrometry. J. Am. Chem. Soc. 136, 12702-12711 (2014)]. Here, we show that the solvent-free intermediates refold into the all- cis PPI helix with high (>90%) efficiency. Moreover, in the absence of solvent, each intermediate appears to utilize the same small set of pathways observed for the solution-phase PPII → PPI transition upon immersion of PPIIaq in 1-propanol. That folding in solution (under conditions where water is displaced by propanol) and folding in vacuo (where energy required for folding is provided by collisional activation) occur along the same pathway is remarkable. Implicit in this statement is that 1-propanol mimics a "dry" environment, similar to the gas phase. We note that intermediates with structures that are similar to PPIIaq can form PPII under the most gentle activation conditions—indicating that some transitions observed in water (i.e. , "we t" folding, are accessible (albeit inefficient) in vacuo. Lastly, these "dry" folding experiments show that PPI (all cis) is favored under "dry" conditions, which underscores the role of water as the major factor promoting preference for trans proline.

  11. Folding of non-Euclidean curved shells

    NASA Astrophysics Data System (ADS)

    Bende, Nakul; Evans, Arthur; Innes-Gold, Sarah; Marin, Luis; Cohen, Itai; Santangelo, Christian; Hayward, Ryan

    2015-03-01

    Origami-based folding of 2D sheets has been of recent interest for a variety of applications ranging from deployable structures to self-folding robots. Though folding of planar sheets follows well-established principles, folding of curved shells involves an added level of complexity due to the inherent influence of curvature on mechanics. In this study, we use principles from differential geometry and thin shell mechanics to establish fundamental rules that govern folding of prototypical creased shells. In particular, we show how the normal curvature of a crease line controls whether the deformation is smooth or discontinuous, and investigate the influence of shell thickness and boundary conditions. We show that snap-folding of shells provides a route to rapid actuation on time-scales dictated by the speed of sound. The simple geometric design principles developed can be applied at any length-scale, offering potential for bio-inspired soft actuators for tunable optics, microfluidics, and robotics. This work was funded by the National Science Foundation through EFRI ODISSEI-1240441 with additional support to S.I.-G. through the UMass MRSEC DMR-0820506 REU program.

  12. Single-molecule observation of protein folding in symmetric GroEL-(GroES)2 complexes.

    PubMed

    Takei, Yodai; Iizuka, Ryo; Ueno, Taro; Funatsu, Takashi

    2012-11-30

    The chaperonin, GroEL, is an essential molecular chaperone that mediates protein folding together with its cofactor, GroES, in Escherichia coli. It is widely believed that the two rings of GroEL alternate between the folding active state coupled to GroES binding during the reaction cycle. In other words, an asymmetric GroEL-GroES complex (the bullet-shaped complex) is formed throughout the cycle, whereas a symmetric GroEL-(GroES)(2) complex (the football-shaped complex) is not formed. We have recently shown that the football-shaped complex coexists with the bullet-shaped complex during the reaction cycle. However, how protein folding proceeds in the football-shaped complex remains poorly understood. Here, we used GFP as a substrate to visualize protein folding in the football-shaped complex by single-molecule fluorescence techniques. We directly showed that GFP folding occurs in both rings of the football-shaped complex. Remarkably, the folding was a sequential two-step reaction, and the kinetics were in excellent agreement with those in the bullet-shaped complex. These results demonstrate that the same reactions take place independently in both rings of the football-shaped complex to facilitate protein folding. PMID:23048033

  13. Can Local Probes Go Global? A Joint Experiment-Simulation Analysis of λ6-85 Folding.

    PubMed

    Sukenik, Shahar; Pogorelov, Taras V; Gruebele, Martin

    2016-06-01

    The process of protein folding is known to involve global motions in a cooperative affair; the structure of most of the protein sequences is gained or lost over a narrow range of temperature, denaturant, or pressure perturbations. At the same time, recent simulations and experiments reveal a complex structural landscape with a rich set of local motions and conformational changes. We couple experimental kinetic and thermodynamic measurements with specifically tailored analysis of simulation data to isolate local versus global folding probes. We find that local probes exhibit lower melting temperatures, smaller surface area changes, and faster kinetics compared to global ones. We also see that certain local probes of folding match the global behavior more closely than others. Our work highlights the importance of using multiple probes to fully characterize protein folding dynamics by theory and experiment. PMID:27101436

  14. Can Local Probes Go Global? A Joint Experiment-Simulation Analysis of λ6-85 Folding.

    PubMed

    Sukenik, Shahar; Pogorelov, Taras V; Gruebele, Martin

    2016-06-01

    The process of protein folding is known to involve global motions in a cooperative affair; the structure of most of the protein sequences is gained or lost over a narrow range of temperature, denaturant, or pressure perturbations. At the same time, recent simulations and experiments reveal a complex structural landscape with a rich set of local motions and conformational changes. We couple experimental kinetic and thermodynamic measurements with specifically tailored analysis of simulation data to isolate local versus global folding probes. We find that local probes exhibit lower melting temperatures, smaller surface area changes, and faster kinetics compared to global ones. We also see that certain local probes of folding match the global behavior more closely than others. Our work highlights the importance of using multiple probes to fully characterize protein folding dynamics by theory and experiment.

  15. The earliest events in protein folding: Helix dynamics in proteins and model peptides

    SciTech Connect

    Dyer, R.B.; Williams, S.; Woodruff, W.H.

    1996-12-31

    The earliest events in protein folding are critically important in determining the folding pathway, but have proved difficult to study by conventional approaches. We have developed new rapid initiation methods and structure-specific probes to interrogate the earliest events of protein folding. Our focus is the pathways. Folding or unfolding reactions are initiated on a fast timescale (10 ns) using a laser induced temperature jump (15 C) and probed with time-resolved infrared spectroscopy. We obtained the kinetics of the helix-coil transition for a model 21-residue peptide. The observed rate constant k{sub obs} = k{sub f} + k{sub u} for reversible kinetics; from the observed rate (6 x 10{sup 6} s{sup -1}) and the equilibrium constant favoring folding of 7.5 at 27 C, we calculate a folding lifetime of 180 ns and an unfolding lifetime of 1.4 {mu}s. The {open_quotes}molten globule{close_quotes} form of apomyoglobin (horse, pH*3, 0.15M NaCl) shows similar kinetics for helix that is unconstrained by tertiary structure (helix with an unusually low Amide I frequency, near 1633 cm{sup -1}). In {open_quotes}native{close_quotes} apomyoglobin (horse, pH*5.3, 10 mM NaCl) two very different rates (45 ns and 70 {mu}s) are observed and we infer that a third occurs on a timescales inaccessible to our experiment (> 1 ms). We suggest that the slower processes are due to helix formation that is rate-limited by the formation of tertiary structure.

  16. Microsecond acquisition of heterogeneous structure in the folding of a TIM barrel protein

    PubMed Central

    Wu, Ying; Kondrashkina, Elena; Kayatekin, Can; Matthews, C. Robert; Bilsel, Osman

    2008-01-01

    The earliest kinetic folding events for (βα)8 barrels reflect the appearance of off-pathway intermediates. Continuous-flow microchannel mixing methods interfaced to small-angle x-ray scattering (SAXS), circular dichroism (CD), time-resolved Förster resonant energy transfer (trFRET), and time-resolved fluorescence anisotropy (trFLAN) have been used to directly monitor global and specific dimensional properties of the partially folded state in the microsecond time range for a representative (βα)8 barrel protein. Within 150 μs, the α-subunit of Trp synthase (αTS) experiences a global collapse and the partial formation of secondary structure. The time resolution of the folding reaction was enhanced with trFRET and trFLAN to show that, within 30 μs, a distinct and autonomous partially collapsed structure has already formed in the N-terminal and central regions but not in the C-terminal region. A distance distribution analysis of the trFRET data confirmed the presence of a heterogeneous ensemble that persists for several hundreds of microseconds. Ready access to locally folded, stable substructures may be a hallmark of repeat-module proteins and the source of early kinetic traps in these very common motifs. Their folding free-energy landscapes should be elaborated to capture this source of frustration. PMID:18757725

  17. Cytochrome c folds through foldon-dependent native-like intermediates in an ordered pathway

    PubMed Central

    Hu, Wenbing; Kan, Zhong-Yuan; Mayne, Leland; Englander, S. Walter

    2016-01-01

    Previous hydrogen exchange (HX) studies of the spontaneous reversible unfolding of Cytochrome c (Cyt c) under native conditions have led to the following conclusions. Native Cyt c (104 residues) is composed of five cooperative folding units, called foldons. The high-energy landscape is dominated by an energy ladder of partially folded forms that differ from each other by one cooperative foldon unit. The reversible equilibrium unfolding of native Cyt c steps up through these intermediate forms to the unfolded state in an energy-ordered sequence, one foldon unit at a time. To more directly study Cyt c intermediates and pathways during normal energetically downhill kinetic folding, the present work used HX pulse labeling analyzed by a fragment separation–mass spectrometry method. The results show that 95% or more of the Cyt c population folds by stepping down through the same set of foldon-dependent pathway intermediates as in energetically uphill equilibrium unfolding. These results add to growing evidence that proteins fold through a classical pathway sequence of native-like intermediates rather than through a vast number of undefinable intermediates and pathways. The present results also emphasize the condition-dependent nature of kinetic barriers, which, with less informative experimental methods (fluorescence, etc.), are often confused with variability in intermediates and pathways. PMID:26966231

  18. New approach to folding with the Coulomb wave function

    SciTech Connect

    Blokhintsev, L. D.; Savin, D. A.; Kadyrov, A. S.; Mukhamedzhanov, A. M.

    2015-05-15

    Due to the long-range character of the Coulomb interaction theoretical description of low-energy nuclear reactions with charged particles still remains a formidable task. One way of dealing with the problem in an integral-equation approach is to employ a screened Coulomb potential. A general approach without screening requires folding of kernels of the integral equations with the Coulomb wave. A new method of folding a function with the Coulomb partial waves is presented. The partial-wave Coulomb function both in the configuration and momentum representations is written in the form of separable series. Each term of the series is represented as a product of a factor depending only on the Coulomb parameter and a function depending on the spatial variable in the configuration space and the momentum variable if the momentum representation is used. Using a trial function, the method is demonstrated to be efficient and reliable.

  19. Effects of confinement on protein folding and protein stability

    NASA Astrophysics Data System (ADS)

    Ping, G.; Yuan, J. M.; Vallieres, M.; Dong, H.; Sun, Z.; Wei, Y.; Li, F. Y.; Lin, S. H.

    2003-05-01

    In a cell, proteins exist in crowded environments; these environments influence their stability and dynamics. Similarly, for an enzyme molecule encapsulated in an inorganic cavity as in biosensors or biocatalysts, confinement and even surface effects play important roles in its stability and dynamics. Using a minimalist model (two-dimensional HP lattice model), we have carried out Monte Carlo simulations to study confinement effects on protein stability. We have calculated heat capacity as a function of temperature using the histogram method and results obtained show that confinement tends to stabilize the folded conformations, consistent with experimental results (some reported here) and previous theoretical analyses. Furthermore, for a protein molecule tethered to a solid surface the stabilization effect can be even greater. We have also investigated the effects of confinement on the kinetics of the refolding and unfolding processes as functions of temperature and box size. As expected, unfolding time increases as box size decreases, however, confinement affects folding times in a more complicated way. Our theoretical results agree with our experimentally observed trends that thermal stability of horseradish peroxidase and acid phosphatase, encapsulated in mesoporous silica, increases as the pore size of the silica matrix decreases.

  20. Folding patterns and shape optimization using SMA-based self-folding laminates

    NASA Astrophysics Data System (ADS)

    Peraza-Hernandez, Edwin A.; Frei, Katherine R.; Hartl, Darren J.; Lagoudas, Dimitris C.

    2014-03-01

    Origami engineering, a discipline encompassing the creation of practical three-dimensional structures from two- dimensional entities via folding operations, has the potential to impact multiple fields of manufacturing and design. In some circumstances, it may be practical to have self-folding capabilities instead of creating folds by external manipulations (as in morphing structures in outer space or on the ocean floor). This paper considers the use of a self-folding laminate composite consisting of two outer layers of thermally actuated shape memory alloy (SMA) wire meshes separated by an inner compliant insulating layer. Methods for designing folding patterns and determining temperature fields to obtain desired shapes and behaviors are proposed. Sheets composed of the self-folding laminate are modeled via finite element analysis (FEA) and the proposed methods are implemented to test their capabilities. One method uses a previously developed and freely available software called Freeform Origami for folding pattern design. The second method entails the use of optimization to determine the localized activation temperatures required to obtain desired shapes or to perform specific functions. The proposed methods are demonstrated to be applicable for the design of folding patterns and determination of activation temperatures for the self-folding laminate by showing successful examples of their implementation. This exploratory study provides new tools that can be integrated into the design framework of self-folding origami structures.

  1. Early events in the disulfide-coupled folding of BPTI.

    PubMed Central

    Bulaj, G.; Goldenberg, D. P.

    1999-01-01

    Recent studies of the refolding of reduced bovine pancreatic trypsin inhibitor (BPTI) have shown that a previously unidentified intermediate with a single disulfide is formed much more rapidly than any other one-disulfide species. This intermediate contains a disulfide that is present in the native protein (between Cys14 and 38), but it is thermodynamically less stable than the other two intermediates with single native disulfides. To characterize the role of the [14-38] intermediate and the factors that favor its formation, detailed kinetic and mutational analyses of the early disulfide-formation steps were carried out. The results of these studies indicate that the formation of [14-38] from the fully reduced protein is favored by both local electrostatic effects, which enhance the reactivities of the Cys14 and 38 thiols, and conformational tendencies that are diminished by the addition of urea and are enhanced at lower temperatures. At 25 degrees C and pH 7.3, approximately 35% of the reduced molecules were found to initially form the 14-38 disulfide, but the majority of these molecules then undergo intramolecular rearrangements to generate non-native disulfides, and subsequently the more stable intermediates with native disulfides. Amino acid replacements, other than those involving Cys residues, were generally found to have only small effects on either the rate of forming [14-38] or its thermodynamic stability, even though many of the same substitutions greatly destabilized the native protein and other disulfide-bonded intermediates. In addition, those replacements that did decrease the steady-state concentration of [14-38] did not adversely affect further folding and disulfide formation. These results suggest that the weak and transient interactions that are often detected in unfolded proteins and early folding intermediates may, in some cases, not persist or promote subsequent folding steps. PMID:10493584

  2. Cross folding in southern Bighorn basin

    SciTech Connect

    Gubbels, T.L.

    1986-08-01

    Analysis of Landsat Thematic Mapper imagery coupled with surface structural investigations of well-exposed folds in the southern Bighorn basin have revealed two northwest-trending folds that have been refolded. The eastern boundary of the Owl Creek Mountains is characterized by a well-defined alignment of folds that extend north-northwest from the Owl Creek thrust front. Bridger monocline, Wildhorse Butte anticline, and Red Hole anticline lie along this trend. Initial Laramide folding, probably during latest Cretaceous time, resulted in a single, continuous, north-northwest-trending anticline with a southwestward vergence. This anticline was progressively unfolded from south to north as the Owl Creek Range was thrust southward over the Wind River basin in earliest Eocene time; scissors-like vertical motion along this flexure rotated the axial surface of the early formed Bridger anticline, resulting in a monocline with a reversed vergence (northeastward). Formation of the Thermopolis/East Warm Springs anticline parallel to the north flank of the range accompanied thrusting and effectively refolded the northern end of the Wildhorse Butte anticline along an east-west axis. Faulting of the oversteepened south limb of the Red Hole cross fold was contemporaneous with folding. Cross-cutting fold axes in this area and the Mud Creek area to the west are best explained by a counterclockwise change in stress direction during the latest phase of the Laramide orogeny. Vertical movement along the eastern side of the Owl Creek Range results from differential motion in the hanging wall of the crystalline thrust sheet.

  3. Cellular folding pathway of a metastable serpin.

    PubMed

    Chandrasekhar, Kshama; Ke, Haiping; Wang, Ning; Goodwin, Theresa; Gierasch, Lila M; Gershenson, Anne; Hebert, Daniel N

    2016-06-01

    Although proteins generally fold to their thermodynamically most stable state, some metastable proteins populate higher free energy states. Conformational changes from metastable higher free energy states to lower free energy states with greater stability can then generate the work required to perform physiologically important functions. However, how metastable proteins fold to these higher free energy states in the cell and avoid more stable but inactive conformations is poorly understood. The serpin family of metastable protease inhibitors uses large conformational changes that are downhill in free energy to inhibit target proteases by pulling apart the protease active site. The serpin antithrombin III (ATIII) targets thrombin and other proteases involved in blood coagulation, and ATIII misfolding can thus lead to thrombosis and other diseases. ATIII has three disulfide bonds, two near the N terminus and one near the C terminus. Our studies of ATIII in-cell folding reveal a surprising, biased order of disulfide bond formation, with early formation of the C-terminal disulfide, before formation of the N-terminal disulfides, critical for folding to the active, metastable state. Early folding of the predominantly β-sheet ATIII domain in this two-domain protein constrains the reactive center loop (RCL), which contains the protease-binding site, ensuring that the RCL remains accessible. N-linked glycans and carbohydrate-binding molecular chaperones contribute to the efficient folding and secretion of functional ATIII. The inability of a number of disease-associated ATIII variants to navigate the folding reaction helps to explain their disease phenotypes. PMID:27222580

  4. Estimation of vocal fold plane in 3D CT images for diagnosis of vocal fold abnormalities.

    PubMed

    Hewavitharanage, Sajini; Gubbi, Jayavardhana; Thyagarajan, Dominic; Lau, Ken; Palaniswami, Marimuthu

    2015-01-01

    Vocal folds are the key body structures that are responsible for phonation and regulating air movement into and out of lungs. Various vocal fold disorders may seriously impact the quality of life. When diagnosing vocal fold disorders, CT of the neck is the commonly used imaging method. However, vocal folds do not align with the normal axial plane of a neck and the plane containing vocal cords and arytenoids does vary during phonation. It is therefore important to generate an algorithm for detecting the actual plane containing vocal folds. In this paper, we propose a method to automatically estimate the vocal fold plane using vertebral column and anterior commissure localization. Gray-level thresholding, connected component analysis, rule based segmentation and unsupervised k-means clustering were used in the proposed algorithm. The anterior commissure segmentation method achieved an accuracy of 85%, a good estimate of the expert assessment. PMID:26736949

  5. Saffman-Taylor fingers with kinetic undercooling

    NASA Astrophysics Data System (ADS)

    Gardiner, Bennett P. J.; McCue, Scott W.; Dallaston, Michael C.; Moroney, Timothy J.

    2015-02-01

    The mathematical model of a steadily propagating Saffman-Taylor finger in a Hele-Shaw channel has applications to two-dimensional interacting streamer discharges which are aligned in a periodic array. In the streamer context, the relevant regularization on the interface is not provided by surface tension but instead has been postulated to involve a mechanism equivalent to kinetic undercooling, which acts to penalize high velocities and prevent blow-up of the unregularized solution. Previous asymptotic results for the Hele-Shaw finger problem with kinetic undercooling suggest that for a given value of the kinetic undercooling parameter, there is a discrete set of possible finger shapes, each analytic at the nose and occupying a different fraction of the channel width. In the limit in which the kinetic undercooling parameter vanishes, the fraction for each family approaches 1 /2 , suggesting that this "selection" of 1 /2 by kinetic undercooling is qualitatively similar to the well-known analog with surface tension. We treat the numerical problem of computing these Saffman-Taylor fingers with kinetic undercooling, which turns out to be more subtle than the analog with surface tension, since kinetic undercooling permits finger shapes which are corner-free but not analytic. We provide numerical evidence for the selection mechanism by setting up a problem with both kinetic undercooling and surface tension and numerically taking the limit that the surface tension vanishes.

  6. Computational and theoretical methods for protein folding.

    PubMed

    Compiani, Mario; Capriotti, Emidio

    2013-12-01

    A computational approach is essential whenever the complexity of the process under study is such that direct theoretical or experimental approaches are not viable. This is the case for protein folding, for which a significant amount of data are being collected. This paper reports on the essential role of in silico methods and the unprecedented interplay of computational and theoretical approaches, which is a defining point of the interdisciplinary investigations of the protein folding process. Besides giving an overview of the available computational methods and tools, we argue that computation plays not merely an ancillary role but has a more constructive function in that computational work may precede theory and experiments. More precisely, computation can provide the primary conceptual clues to inspire subsequent theoretical and experimental work even in a case where no preexisting evidence or theoretical frameworks are available. This is cogently manifested in the application of machine learning methods to come to grips with the folding dynamics. These close relationships suggested complementing the review of computational methods within the appropriate theoretical context to provide a self-contained outlook of the basic concepts that have converged into a unified description of folding and have grown in a synergic relationship with their computational counterpart. Finally, the advantages and limitations of current computational methodologies are discussed to show how the smart analysis of large amounts of data and the development of more effective algorithms can improve our understanding of protein folding.

  7. Protein Folding and Mechanisms of Proteostasis

    PubMed Central

    Díaz-Villanueva, José Fernando; Díaz-Molina, Raúl; García-González, Victor

    2015-01-01

    Highly sophisticated mechanisms that modulate protein structure and function, which involve synthesis and degradation, have evolved to maintain cellular homeostasis. Perturbations in these mechanisms can lead to protein dysfunction as well as deleterious cell processes. Therefore in recent years the etiology of a great number of diseases has been attributed to failures in mechanisms that modulate protein structure. Interconnections among metabolic and cell signaling pathways are critical for homeostasis to converge on mechanisms associated with protein folding as well as for the preservation of the native structure of proteins. For instance, imbalances in secretory protein synthesis pathways lead to a condition known as endoplasmic reticulum (ER) stress which elicits the adaptive unfolded protein response (UPR). Therefore, taking this into consideration, a key part of this paper is developed around the protein folding phenomenon, and cellular mechanisms which support this pivotal condition. We provide an overview of chaperone protein function, UPR via, spatial compartmentalization of protein folding, proteasome role, autophagy, as well as the intertwining between these processes. Several diseases are known to have a molecular etiology in the malfunction of mechanisms responsible for protein folding and in the shielding of native structure, phenomena which ultimately lead to misfolded protein accumulation. This review centers on our current knowledge about pathways that modulate protein folding, and cell responses involved in protein homeostasis. PMID:26225966

  8. Experimental investigation of protein folding and misfolding.

    PubMed

    Dobson, Christopher M

    2004-09-01

    Newly synthesised proteins need to fold, often to intricate and close-packed structures, in order to function. The underlying mechanism by which this complex process takes place both in vitro and in vivo is now becoming understood, at least in general terms, as a result of the application of a wide range of biophysical and computational methods used in combination with the techniques of biochemistry and protein engineering. It is increasingly apparent, however, that folding is not only crucial for generating biological activity, but that it is also coupled to a wide range of processes within the cell, ranging from the trafficking of proteins to specific organelles to the regulation of cell growth and differentiation. Not surprisingly, therefore, the failure of proteins to fold appropriately, or to remain correctly folded, is associated with a large number of cellular malfunctions that give rise to disease. Misfolding, and its consequences such as aggregation, can be investigated by extending the types of techniques used to study the normal folding process. Application of these techniques is enabling the development of a unified description of the interconversion and regulation of the different conformational states available to proteins in living systems. Such a description proves a generic basis for understanding the fundamental links between protein misfolding and its associated clinical disorders, such as Alzheimer's disease and Type II diabetes, and for exploring novel therapeutic strategies directed at their prevention and treatment on a rational basis.

  9. A sweet code for glycoprotein folding.

    PubMed

    Caramelo, Julio J; Parodi, Armando J

    2015-11-14

    Glycoprotein synthesis is initiated in the endoplasmic reticulum (ER) lumen upon transfer of a glycan (Glc3Man9GlcNAc2) from a lipid derivative to Asn residues (N-glycosylation). N-Glycan-dependent quality control of glycoprotein folding in the ER prevents exit to Golgi of folding intermediates, irreparably misfolded glycoproteins and incompletely assembled multimeric complexes. It also enhances folding efficiency by preventing aggregation and facilitating formation of proper disulfide bonds. The control mechanism essentially involves four components, resident lectin-chaperones (calnexin and calreticulin) that recognize monoglucosylated polymannose protein-linked glycans, lectin-associated oxidoreductase acting on monoglucosylated glycoproteins (ERp57), a glucosyltransferase that creates monoglucosylated epitopes in protein-linked glycans (UGGT) and a glucosidase (GII) that removes the glucose units added by UGGT. This last enzyme is the only mechanism component sensing glycoprotein conformations as it creates monoglucosylated glycans exclusively in not properly folded glycoproteins or in not completely assembled multimeric glycoprotein complexes. Glycoproteins that fail to properly fold are eventually driven to proteasomal degradation in the cytosol following the ER-associated degradation pathway, in which the extent of N-glycan demannosylation by ER mannosidases play a relevant role in the identification of irreparably misfolded glycoproteins.

  10. Petrofabric test of viscous folding theory

    NASA Astrophysics Data System (ADS)

    Onasch, Charles M.

    1984-06-01

    Compression and extension axes are deduced from quartz deformation lamellae in a quartzite and a graywacke folded into an asymetrical syncline. Deformation lamellae fabrics in the two sandstones are distinctly different. In the graywacke, regardless of bedding orientation or position on the fold, compression axes are normal or nearly normal to the axial planar rough cleavage. Extension axes generally lie in the cleavage plane, parallel to dip. In most quartzite samples, compression axes are parallel or subparallel to bedding, at high angles to the fold axis and extension axes are normal to bedding. Two samples from the very base of the formation indicate compression parallel to the fold axis with extension parallel to bedding, at high angles to the fold axis. One of these two shows both patterns. The lamellae fabric geometry in these two samples suggests the presence of a neutral surface in the quartzite. The lamellae-derived compression and extension axes are in good agreement with the buckling behavior of a viscous layer (quartzite) embedded in a less viscous medium (graywacke and shale below and shale and carbonate above).

  11. Towards a systematic classification of protein folds

    NASA Astrophysics Data System (ADS)

    Lindgård, Per-Anker; Bohr, Henrik

    1997-10-01

    A lattice model Hamiltonian is suggested for protein structures that can explain the division into structural fold classes during the folding process. Proteins are described by chains of secondary structure elements, with the hinges in between being the important degrees of freedom. The protein structures are given a unique name, which simultaneously represent a linear string of physical coupling constants describing hinge spin interactions. We have defined a metric and a precise distance measure between the fold classes. An automated procedure is constructed in which any protein structure in the usual protein data base coordinate format can be transformed into the proposed chain representation. Taking into account hydrophobic forces we have found a mechanism for the formation of domains with a unique fold containing predicted magic numbers \\{4,6,9,12,16,18,...\\} of secondary structures and multiples of these domains. It is shown that the same magic numbers are robust and occur as well for packing on other nonclosed packed lattices. We have performed a statistical analysis of available protein structures and found agreement with the predicted preferred abundances of proteins with a predicted magic number of secondary structures. Thermodynamic arguments for the increased abundance and a phase diagram for the folding scenario are given. This includes an intermediate high symmetry phase, the parent structures, between the molten globule and the native states. We have made an exhaustive enumeration of dense lattice animals on a cubic lattice for acceptance number Z=4 and Z=5 up to 36 vertices.

  12. Chemical kinetics and combustion modeling

    SciTech Connect

    Miller, J.A.

    1993-12-01

    The goal of this program is to gain qualitative insight into how pollutants are formed in combustion systems and to develop quantitative mathematical models to predict their formation rates. The approach is an integrated one, combining low-pressure flame experiments, chemical kinetics modeling, theory, and kinetics experiments to gain as clear a picture as possible of the process in question. These efforts are focused on problems involved with the nitrogen chemistry of combustion systems and on the formation of soot and PAH in flames.

  13. Parameters affecting in vitro oxidation/folding of maurotoxin, a four-disulphide-bridged scorpion toxin.

    PubMed Central

    di Luccio, E; Azulay, D O; Regaya, I; Fajloun, Z; Sandoz, G; Mansuelle, P; Kharrat, R; Fathallah, M; Carrega, L; Estève, E; Rochat, H; De Waard, M; Sabatier, J M

    2001-01-01

    Maurotoxin (MTX) is a 34-mer scorpion toxin cross-linked by four disulphide bridges that acts on various K(+) channel subtypes. MTX adopts a disulphide bridge organization of the type C1-C5, C2-C6, C3-C4 and C7-C8, and folds according to the common alpha/beta scaffold reported for other known scorpion toxins. Here we have investigated the process and kinetics of the in vitro oxidation/folding of reduced synthetic L-MTX (L-sMTX, where L-MTX contains only L-amino acid residues). During the oxidation/folding of reduced L-sMTX, the oxidation intermediates were blocked by iodoacetamide alkylation of free cysteine residues, and analysed by MS. The L-sMTX intermediates appeared sequentially over time from the least (intermediates with one disulphide bridge) to the most oxidized species (native-like, four-disulphide-bridged L-sMTX). The mathematical formulation of the diffusion-collision model being inadequate to accurately describe the kinetics of oxidation/folding of L-sMTX, we have formulated a derived mathematical description that better fits the experimental data. Using this mathematical description, we have compared for the first time the oxidation/folding of L-sMTX with that of D-sMTX, its stereoisomer that contains only D-amino acid residues. Several experimental parameters, likely to affect the oxidation/folding process, were studied further; these included temperature, pH, ionic strength, redox potential and concentration of reduced toxin. We also assessed the effects of some cellular enzymes, peptidylprolyl cis-trans isomerase (PPIase) and protein disulphide isomerase (PDI), on the folding pathways of reduced L-sMTX and D-sMTX. All the parameters tested affect the oxidative folding of sMTX, and the kinetics of this process were indistinguishable for L-sMTX and D-sMTX, except when stereospecific enzymes were used. The most efficient conditions were found to be: 50 mM Tris/HCl/1.4 mM EDTA, pH 7.5, supplemented by 0.5 mM PPIase and 50 units/ml PDI for 0.1 m

  14. Proteopedia: Rossmann Fold: A Beta-Alpha-Beta Fold at Dinucleotide Binding Sites

    ERIC Educational Resources Information Center

    Hanukoglu, Israel

    2015-01-01

    The Rossmann fold is one of the most common and widely distributed super-secondary structures. It is composed of a series of alternating beta strand (ß) and alpha helical (a) segments wherein the ß-strands are hydrogen bonded forming a ß-sheet. The initial beta-alpha-beta (ßaß) fold is the most conserved segment of Rossmann folds. As this segment…

  15. Molecular Simulations of Cotranslational Protein Folding: Fragment Stabilities, Folding Cooperativity, and Trapping in the Ribosome

    PubMed Central

    Elcock, Adrian H

    2006-01-01

    Although molecular simulation methods have yielded valuable insights into mechanistic aspects of protein refolding in vitro, they have up to now not been used to model the folding of proteins as they are actually synthesized by the ribosome. To address this issue, we report here simulation studies of three model proteins: chymotrypsin inhibitor 2 (CI2), barnase, and Semliki forest virus protein (SFVP), and directly compare their folding during ribosome-mediated synthesis with their refolding from random, denatured conformations. To calibrate the methodology, simulations are first compared with in vitro data on the folding stabilities of N-terminal fragments of CI2 and barnase; the simulations reproduce the fact that both the stability and thermal folding cooperativity increase as fragments increase in length. Coupled simulations of synthesis and folding for the same two proteins are then described, showing that both fold essentially post-translationally, with mechanisms effectively identical to those for refolding. In both cases, confinement of the nascent polypeptide chain within the ribosome tunnel does not appear to promote significant formation of native structure during synthesis; there are however clear indications that the formation of structure within the nascent chain is sensitive to location within the ribosome tunnel, being subject to both gain and loss as the chain lengthens. Interestingly, simulations in which CI2 is artificially stabilized show a pronounced tendency to become trapped within the tunnel in partially folded conformations: non-cooperative folding, therefore, appears in the simulations to exert a detrimental effect on the rate at which fully folded conformations are formed. Finally, simulations of the two-domain protease module of SFVP, which experimentally folds cotranslationally, indicate that for multi-domain proteins, ribosome-mediated folding may follow different pathways from those taken during refolding. Taken together, these

  16. Non-cylindrical fold growth in the Zagros fold and thrust belt (Kurdistan, NE-Iraq)

    NASA Astrophysics Data System (ADS)

    Bartl, Nikolaus; Bretis, Bernhard; Grasemann, Bernhard; Lockhart, Duncan

    2010-05-01

    The Zagros mountains extends over 1800 km from Kurdistan in N-Iraq to the Strait of Hormuz in Iran and is one of the world most promising regions for the future hydrocarbon exploration. The Zagros Mountains started to form as a result of the collision between the Eurasian and Arabian Plates, whose convergence began in the Late Cretaceous as part of the Alpine-Himalayan orogenic system. Geodetic and seismological data document that both plates are still converging and that the fold and thrust belt of the Zagros is actively growing. Extensive hydrocarbon exploration mainly focuses on the antiforms of this fold and thrust belt and therefore the growth history of the folds is of great importance. This work investigates by means of structural field work and quantitative geomorphological techniques the progressive fold growth of the Permam, Bana Bawi- and Safeen- Anticlines located in the NE of the city of Erbil in the Kurdistan region of Northern Iraq. This part of the Zagros fold and thrust belt belongs to the so-called Simply Folded Belt, which is dominated by gentle to open folding. Faults or fault related folds have only minor importance. The mechanical anisotropy of the formations consisting of a succession of relatively competent (massive dolomite and limestone) and incompetent (claystone and siltstone) sediments essentially controls the deformation pattern with open to gentle parallel folding of the competent layers and flexural flow folding of the incompetent layers. The characteristic wavelength of the fold trains is around 10 km. Due to faster erosion of the softer rock layers in the folded sequence, the more competent lithologies form sharp ridges with steeply sloping sides along the eroded flanks of the anticlines. Using an ASTER digital elevation model in combination with geological field data we quantified 250 drainage basins along the different limbs of the subcylindrical Permam, Bana Bawi- and Safeen- Anticlines. Geomorphological indices of the drainage

  17. Influence of the ventricular folds on a voice source with specified vocal fold motion1

    PubMed Central

    McGowan, Richard S.; Howe, Michael S.

    2010-01-01

    The unsteady drag on the vocal folds is the major source of sound during voiced speech. The drag force is caused by vortex shedding from the vocal folds. The influence of the ventricular folds (i.e., the “false” vocal folds that protrude into the vocal tract a short distance downstream of the glottis) on the drag and the voice source are examined in this paper by means of a theoretical model involving vortex sheets in a two-dimensional geometry. The effect of the ventricular folds on the output acoustic pressure is found to be small when the movement of the vocal folds is prescribed. It is argued that the effect remains small when fluid-structure interactions account for vocal fold movement. These conclusions can be justified mathematically when the characteristic time scale for change in the velocity of the glottal jet is large compared to the time it takes for a vortex disturbance to be convected through the vocal fold and ventricular fold region. PMID:20329852

  18. Energetic frustrations in protein folding at residue resolution: a homologous simulation study of Im9 proteins.

    PubMed

    Sun, Yunxiang; Ming, Dengming

    2014-01-01

    Energetic frustration is becoming an important topic for understanding the mechanisms of protein folding, which is a long-standing big biological problem usually investigated by the free energy landscape theory. Despite the significant advances in probing the effects of folding frustrations on the overall features of protein folding pathways and folding intermediates, detailed characterizations of folding frustrations at an atomic or residue level are still lacking. In addition, how and to what extent folding frustrations interact with protein topology in determining folding mechanisms remains unclear. In this paper, we tried to understand energetic frustrations in the context of protein topology structures or native-contact networks by comparing the energetic frustrations of five homologous Im9 alpha-helix proteins that share very similar topology structures but have a single hydrophilic-to-hydrophobic mutual mutation. The folding simulations were performed using a coarse-grained Gō-like model, while non-native hydrophobic interactions were introduced as energetic frustrations using a Lennard-Jones potential function. Energetic frustrations were then examined at residue level based on φ-value analyses of the transition state ensemble structures and mapped back to native-contact networks. Our calculations show that energetic frustrations have highly heterogeneous influences on the folding of the four helices of the examined structures depending on the local environment of the frustration centers. Also, the closer the introduced frustration is to the center of the native-contact network, the larger the changes in the protein folding. Our findings add a new dimension to the understanding of protein folding the topology determination in that energetic frustrations works closely with native-contact networks to affect the protein folding.

  19. The Role of Electrostatic Interactions in Folding of β-Proteins

    PubMed Central

    Davis, Caitlin M.; Dyer, R. Brian

    2016-01-01

    Atomic-level molecular dynamic simulations are capable of fully folding structurally diverse proteins; however, they are limited in their ability to accurately represent electrostatic interactions. Here we have experimentally tested the role of charged residues on stability and folding kinetics of one of the most widely simulated β-proteins, the WW domain. The folding of wild type Pin1 WW domain, which has two positively charged residues in the first turn, was compared to the fast folding mutant FiP35 Pin1, which introduces a negative charge into the first turn. A combination of FTIR spectroscopy and laser-induced temperature-jump coupled with infrared spectroscopy was used to probe changes in the amide I region. The relaxation dynamics of the peptide backbone, β-sheets and β-turns, and negatively charged aspartic acid side chain of FiP35 were measured independently by probing the corresponding bands assigned in the amide I region. Folding is initiated in the turns and the β-sheets form last. While the global folding mechanism is in good agreement with simulation predictions, we observe changes in the protonation state of aspartic acid during folding that have not been captured by simulation methods. The protonation state of aspartic acid is coupled to protein folding; the apparent pKa of aspartic acid in the folded protein is 6.4. The dynamics of the aspartic acid follow the dynamics of the intermediate phase, supporting assignment of this phase to formation of the first hairpin. These results demonstrate the importance of electrostatic interactions in turn stability and formation of extended β-sheet structures. PMID:26750867

  20. Thermal stability of idealized folded carbyne loops

    PubMed Central

    2013-01-01

    Self-unfolding items provide a practical convenience, wherein ring-like frames are contorted into a state of equilibrium and subsequently  pop up’ or deploy when perturbed from a folded structure. Can the same process be exploited at the molecular scale? At the limiting scale is a closed chain of single atoms, used here to investigate the limits of stability of such folded ring structures via full atomistic molecular dynamics. Carbyne is a one-dimensional carbon allotrope composed of sp-hybridized carbon atoms. Here, we explore the stability of idealized carbyne loops as a function of chain length, curvature, and temperature, and delineate an effective phase diagram between folded and unfolded states. We find that while overall curvature is reduced, in addition to torsional and self-adhesive energy barriers, a local increase in curvature results in the largest impedance to unfolding. PMID:24252156

  1. Microbial Manipulation of the Amyloid Fold

    PubMed Central

    DePas, William H.

    2012-01-01

    Microbial biofilms are encased in a protein, DNA and polysaccharide matrix that protects the community, promotes interactions with the environment, and helps cells to adhere together. The protein component of these matrices is often a remarkably stable, β-sheet-rich polymer called amyloid. Amyloids form ordered, self-templating fibers that are highly aggregative, making them a valuable biofilm component. Some eukaryotic proteins inappropriately adopt the amyloid fold and these misfolded protein aggregates disrupt normal cellular proteostasis, which can cause significant cytotoxicity. Indeed, until recently amyloids were considered solely the result of protein misfolding. However, research over the past decade has revealed how various organisms have capitalized on the amyloid fold by developing sophisticated biogenesis pathways that coordinate gene expression, protein folding, and secretion so that amyloid-related toxicities are minimized. How microbes manipulate amyloids, by augmenting their advantageous properties and by reducing their undesirable properties, will be the subject of this review. PMID:23108148

  2. Transversal Clifford gates on folded surface codes

    NASA Astrophysics Data System (ADS)

    Moussa, Jonathan E.

    2016-10-01

    Surface and color codes are two forms of topological quantum error correction in two spatial dimensions with complementary properties. Surface codes have lower-depth error detection circuits and well-developed decoders to interpret and correct errors, while color codes have transversal Clifford gates and better code efficiency in the number of physical qubits needed to achieve a given code distance. A formal equivalence exists between color codes and folded surface codes, but it does not guarantee the transferability of any of these favorable properties. However, the equivalence does imply the existence of constant-depth circuit implementations of logical Clifford gates on folded surface codes. We achieve and improve this result by constructing two families of folded surface codes with transversal Clifford gates. This construction is presented generally for qudits of any dimension. The specific application of these codes to universal quantum computation based on qubit fusion is also discussed.

  3. Stretching and folding in finite time.

    PubMed

    Ma, Tian; Ouellette, Nicholas T; Bollt, Erik M

    2016-02-01

    Complex flows mix efficiently, and this process can be understood by considering the stretching and folding of material volumes. Although many metrics have been devised to characterize stretching, fewer are able to capture folding in a quantitative way in spatiotemporally variable flows. Here, we extend our previous methods based on the finite-time curving of fluid-element trajectories to nonzero scales and show that this finite-scale finite-time curvature contains information about both stretching and folding. We compare this metric to the more commonly used finite-time Lyapunov exponent and illustrate our methods using experimental flow-field data from a quasi-two-dimensional laboratory flow. Our new analysis tools add to the growing set of Lagrangian methods for characterizing mixing in complex, aperiodic fluid flows.

  4. Exact folded-band chaotic oscillator

    NASA Astrophysics Data System (ADS)

    Corron, Ned J.; Blakely, Jonathan N.

    2012-06-01

    An exactly solvable chaotic oscillator with folded-band dynamics is shown. The oscillator is a hybrid dynamical system containing a linear ordinary differential equation and a nonlinear switching condition. Bounded oscillations are provably chaotic, and successive waveform maxima yield a one-dimensional piecewise-linear return map with segments of both positive and negative slopes. Continuous-time dynamics exhibit a folded-band topology similar to Rössler's oscillator. An exact solution is written as a linear convolution of a fixed basis pulse and a discrete binary sequence, from which an equivalent symbolic dynamics is obtained. The folded-band topology is shown to be dependent on the symbol grammar.

  5. Inclusion Kinetics of Polyrotaxanes

    NASA Astrophysics Data System (ADS)

    Yokoyama, Hideaki; Takahashi, Shoko; Ito, Kohzo; Yamada, Norifumi

    Inclusion complex (IC) formation of α-cyclodextrin (α-CD) and poly(ethylene glycol) (PEG) brush in water was investigated by Surface Plasmon Resonance Spectroscopy(SPR), neutron reflectometry(NR) and grazing incident wide angle X-ray scattering(GISANS). Spontaneous IC formation of α-CD with PEG (polyrotaxanes) is believed to be due to hydrophobic interaction between the hydrophobic interior of α-CD and PEG; however, the detail of the IC formation kinetics has not been observed because IC formation results in aggregation and precipitation of the complex. SPR revealed that IC formation occurs after induction period, which often appears in crystallization. When concentration of α-CD solution is 10%, IC consisting randomly oriented α-CD polycrystal appeared. In contrast, when the concentration of α-CD solution is 5%, a uniform 10-nm-thick IC layer with α-CD stacked perpendicular to the substrate appeared. 10-nm-thick IC was also found in the diluted PEG brush in contact with a 10% α-CD solution. The characteristic 10-nm-thick layer is related to the folded crystalline structure of α-CD on PEG brush. Such crystallization was proved to be the main driving force for IC formation.

  6. Ontogeny of the mouse vocal fold epithelium

    PubMed Central

    Lungova, Vlasta; Verheyden, Jamie M.; Herriges, John; Sun, Xin; Thibeault, Susan L.

    2015-01-01

    This investigation provides the first systematic determination of the cellular and molecular progression of vocal fold (VF) epithelium development in a murine model. We define five principal developmental events that constitute the progression from VF initiation in the embryonic anterior foregut tube to fully differentiated and functional adult tissue. These developmental events include (1) the initiation of the larynx and vocal folds with apposition of the lateral walls of the primitive laryngopharynx (embryonic (E) day 10.5); (2) the establishment of the epithelial lamina with fusion of the lateral walls of the primitive laryngopharynx (E11.5); (3) the epithelial lamina recanalization and separation of VFs (E13.5–18.5); (4) the stratification of the vocal folds (E13.5–18.5); and (5) the maturation of vocal fold epithelium (postnatal stages). The illustration of these morphogenetic events is substantiated by dynamic changes in cell proliferation and apoptosis, as well as the expression pattern of key transcription factors, FOXA2, SOX2 and NKX2-1 that specify and pattern the foregut endoderm. Furthermore, we documented the gradual conversion of VF epithelial cells from simple precursors expressing cytokeratins 8 and 18 in the embryo into mature stratified epithelial cells also expressing cytokeratins 5 and 14 in the adult. Interestingly, in the adult, cytokeratins 5 and 14 appear to be expressed in all cell layers in the VF, in contrast to their preferential localization to the basal cell layer in surrounding epithelium. To begin investigating the role of signaling molecules in vocal fold development, we characterized the expression pattern of SHH pathway genes, and how loss of Shh affects vocal fold development in the mutant. This study defines the cellular and molecular context and serves as the necessary foundation for future functional investigations of VF formation. PMID:25601450

  7. Circular permutant GFP insertion folding reporters

    DOEpatents

    Waldo, Geoffrey S.; Cabantous, Stephanie

    2008-06-24

    Provided are methods of assaying and improving protein folding using circular permutants of fluorescent proteins, including circular permutants of GFP variants and combinations thereof. The invention further provides various nucleic acid molecules and vectors incorporating such nucleic acid molecules, comprising polynucleotides encoding fluorescent protein circular permutants derived from superfolder GFP, which polynucleotides include an internal cloning site into which a heterologous polynucleotide may be inserted in-frame with the circular permutant coding sequence, and which when expressed are capable of reporting on the degree to which a polypeptide encoded by such an inserted heterologous polynucleotide is correctly folded by correlation with the degree of fluorescence exhibited.

  8. Circular permutant GFP insertion folding reporters

    SciTech Connect

    Waldo, Geoffrey S.; Cabantous, Stephanie

    2013-04-16

    Provided are methods of assaying and improving protein folding using circular permutants of fluorescent proteins, including circular permutants of GFP variants and combinations thereof. The invention further provides various nucleic acid molecules and vectors incorporating such nucleic acid molecules, comprising polynucleotides encoding fluorescent protein circular permutants derived from superfolder GFP, which polynucleotides include an internal cloning site into which a heterologous polynucleotide may be inserted in-frame with the circular permutant coding sequence, and which when expressed are capable of reporting on the degree to which a polypeptide encoded by such an inserted heterologous polynucleotide is correctly folded by correlation with the degree of fluorescence exhibited.

  9. Circular permutant GFP insertion folding reporters

    SciTech Connect

    Waldo, Geoffrey S; Cabantous, Stephanie

    2013-02-12

    Provided are methods of assaying and improving protein folding using circular permutants of fluorescent proteins, including circular permutants of GFP variants and combinations thereof. The invention further provides various nucleic acid molecules and vectors incorporating such nucleic acid molecules, comprising polynucleotides encoding fluorescent protein circular permutants derived from superfolder GFP, which polynucleotides include an internal cloning site into which a heterologous polynucleotide may be inserted in-frame with the circular permutant coding sequence, and which when expressed are capable of reporting on the degree to which a polypeptide encoded by such an inserted heterologous polynucleotide is correctly folded by correlation with the degree of fluorescence exhibited.

  10. Circular permutant GFP insertion folding reporters

    SciTech Connect

    Waldo, Geoffrey S.; Cabantous, Stephanie

    2011-06-14

    Provided are methods of assaying and improving protein folding using circular permutants of fluorescent proteins, including circular permutants of GFP variants and combinations thereof. The invention further provides various nucleic acid molecules and vectors incorporating such nucleic acid molecules, comprising polynucleotides encoding fluorescent protein circular permutants derived from superfolder GFP, which polynucleotides include an internal cloning site into which a heterologous polynucleotide may be inserted in-frame with the circular permutant coding sequence, and which when expressed are capable of reporting on the degree to which a polypeptide encoded by such an inserted heterologous polynucleotide is correctly folded by correlation with the degree of fluorescence exhibited.

  11. Control of folding by gravity and matrix thickness: Implications for large-scale folding

    NASA Astrophysics Data System (ADS)

    Schmalholz, S. M.; Podladchikov, Y. Y.; Burg, J.-P.

    2002-01-01

    We show that folding of a non-Newtonian layer resting on a homogeneous Newtonian matrix with finite thickness under influence of gravity can occur by three modes: (1) matrix-controlled folding, dependent on the effective viscosity contrast between layer and matrix, (2) gravity-controlled folding, dependent on the Argand number (the ratio of the stress caused by gravity to the stress caused by shortening), and (3) detachment folding, dependent on the ratio of matrix thickness to layer thickness. We construct a phase diagram that defines the transitions between each of the three folding modes. Our priority is transparency of the analytical derivations (e.g., thin-plate versus thick-plate approximations), which permits complete classification of the folding modes involving a minimum number of dimensionless parameters. Accuracy and sensitivity of the analytical results to model assumptions are investigated. In particular, depth dependence of matrix rheology is only important for folding over a narrow range of material parameters. In contrast, strong depth dependence of the viscosity of the folding layer limits applicability of ductile rheology and leads to a viscoelastic transition. Our theory is applied to estimate the effective thickness of the folded central Asian upper crust using the ratio of topographic wavelength to Moho depth. Phase diagrams based on geometrical parameters show that gravity does not significantly control folding in the Jura and the Zagros Mountains but does control folding in central Asia. Applicability conditions of viscous and thin sheet models for large-scale lithospheric deformation, derived in terms of the Argand number, have implications for the plate-like style of planetary tectonics.

  12. MICROFLUIDIC MIXERS FOR THE INVESTIGATION OF PROTEIN FOLDING USING SYNCHROTRON RADIATION CIRCULAR DICHROISM SPECTROSCOPY

    SciTech Connect

    Kane, A; Hertzog, D; Baumgartel, P; Lengefeld, J; Horsley, D; Schuler, B; Bakajin, O

    2006-03-20

    The purpose of this study is to design, fabricate and optimize microfluidic mixers to investigate the kinetics of protein secondary structure formation with Synchrotron Radiation Circular Dichroism (SRCD) spectroscopy. The mixers are designed to rapidly initiate protein folding reaction through the dilution of denaturant. The devices are fabricated out of fused silica, so that they are transparent in the UV. We present characterization of mixing in the fabricated devices, as well as the initial SRCD data on proteins inside the mixers.

  13. Designing the structure and folding pathway of modular topological bionanostructures.

    PubMed

    Ljubetič, A; Drobnak, I; Gradišar, H; Jerala, R

    2016-04-18

    Polypeptides and polynucleotides are programmable natural polymers whose linear sequence can be easily designed and synthesized by the cellular transcription/translation machinery. Nature primarily uses proteins as the molecular machines and nucleic acids as the medium for the manipulation of heritable information. A protein's tertiary structure and function is defined by multiple cooperative weak long-range interactions that have been optimized through evolution. DNA nanotechnology uses orthogonal pairwise interacting modules of complementary nucleic acids as a strategy to construct defined complex 3D structures. A similar approach has recently been applied to protein design, using orthogonal dimerizing coiled-coil segments as interacting modules. When concatenated into a single polypeptide chain, they self-assemble into the 3D structure defined by the topology of interacting modules within the chain. This approach allows the construction of geometric polypeptide scaffolds, bypassing the folding problem of compact proteins by relying on decoupled pairwise interactions. However, the folding pathway still needs to be optimized in order to allow rapid self-assembly under physiological conditions. Again the modularity of designed topological structures can be used to define the rules that guide the folding pathway of long polymers, such as DNA, based on the stability and topology of connected building modules. This approach opens the way towards incorporation of designed foldamers in biological systems and their functionalization. PMID:27001947

  14. Filling Up the Heme Pocket Stabilizes Apomyoglobin and Speeds Up Its Folding

    PubMed Central

    2015-01-01

    Wild type apomyoglobin folds in at least two steps: the ABGH core rapidly, followed much later by the heme-binding CDEF core. We hypothesize that the evolved heme-binding function of the CDEF core frustrates its folding: it has a smaller contact order and is no more complex topologically than ABGH, and thus, it should be able to fold faster. Therefore, filling up the empty heme cavity of apomyoglobin with larger, hydrophobic side chains should significantly stabilize the protein and increase its folding rate. Molecular dynamics simulations allowed us to design four different mutants with bulkier side chains that increase the native bias of the CDEF region. In vitro thermal denaturation shows that the mutations increase folding stability and bring the protein closer to two-state behavior, as judged by the difference of fluorescence- and circular dichroism-detected protein stability. Millisecond stopped flow measurements of the mutants exhibit refolding kinetics that are over 4 times faster than the wild type’s. We propose that myoglobin-like proteins not evolved to bind heme are equally stable, and find an example. Our results illustrate how evolution for function can force proteins to adapt frustrated folding mechanisms, despite having simple topologies. PMID:24456280

  15. Filling up the heme pocket stabilizes apomyoglobin and speeds up its folding.

    PubMed

    Goodman, J S; Chao, S-H; Pogorelov, T V; Gruebele, M

    2014-06-19

    Wild type apomyoglobin folds in at least two steps: the ABGH core rapidly, followed much later by the heme-binding CDEF core. We hypothesize that the evolved heme-binding function of the CDEF core frustrates its folding: it has a smaller contact order and is no more complex topologically than ABGH, and thus, it should be able to fold faster. Therefore, filling up the empty heme cavity of apomyoglobin with larger, hydrophobic side chains should significantly stabilize the protein and increase its folding rate. Molecular dynamics simulations allowed us to design four different mutants with bulkier side chains that increase the native bias of the CDEF region. In vitro thermal denaturation shows that the mutations increase folding stability and bring the protein closer to two-state behavior, as judged by the difference of fluorescence- and circular dichroism-detected protein stability. Millisecond stopped flow measurements of the mutants exhibit refolding kinetics that are over 4 times faster than the wild type's. We propose that myoglobin-like proteins not evolved to bind heme are equally stable, and find an example. Our results illustrate how evolution for function can force proteins to adapt frustrated folding mechanisms, despite having simple topologies. PMID:24456280

  16. Folding of the Tetrahymena Ribozyme by Polyamines: Importance of Counterion Valence and Size

    SciTech Connect

    Koculi,E.; Lee, N.; Thirumalai, D.; Woodson, S.

    2004-01-01

    Polyamines are abundant metabolites that directly influence gene expression. Although the role of polyamines in DNA condensation is well known, their role in RNA folding is less understood. Non-denaturing gel electrophoresis was used to monitor the equilibrium folding transitions of the Tetrahymena ribozyme in the presence of polyamines. All of the polyamines tested induce near-native structures that readily convert to the native conformation in Mg{sup 2+}. The stability of the folded structure increases with the charge of the polyamine and decreases with the size of the polyamine. When the counterion excluded volume becomes large, the transition to the native state does not go to completion even under favorable folding conditions. Brownian dynamics simulations of a model polyelectrolyte suggest that the kinetics of counterion-mediated collapse and the dimensions of the collapsed RNA chains depend on the structure of the counterion. The results are consistent with delocalized condensation of polyamines around the RNA. However, the effective charge of the counterions is lowered by their excluded volume. The stability of the folded RNA is enhanced when the spacing between amino groups matches the distance between adjacent phosphate groups. These results show how changes in intracellular polyamine concentrations could alter RNA folding pathways.

  17. Folding analysis of the most complex Stevedore’s protein knot

    PubMed Central

    Wang, Iren; Chen, Szu-Yu; Hsu, Shang-Te Danny

    2016-01-01

    DehI is a homodimeric haloacid dehalogenase from Pseudomonas putida that contains the most complex 61 Stevedore’s protein knot within its folding topology. To examine how DehI attains such an intricate knotted topology we combined far-UV circular dichroism (CD), intrinsic fluorescence spectroscopy and small angle X-ray scattering (SAXS) to investigate its folding mechanism. Equilibrium unfolding of DehI by chemical denaturation indicated the presence of two highly populated folding intermediates, I and I’. While the two intermediates vary in secondary structure contents and tertiary packing according to CD and intrinsic fluorescence, respectively, their overall dimension and compactness are similar according to SAXS. Three single-tryptophan variants (W34, W53, and W196) were generated to probe non-cooperative unfolding events localized around the three fluorophores. Kinetic fluorescence measurements indicated that the transition from the intermediate I’ to the unfolded state is rate limiting. Our multiparametric folding analyses suggest that DehI unfolds through a linear folding pathway with two distinct folding intermediates by initial hydrophobic collapse followed by nucleation condensation, and that knotting precedes the formation of secondary structures. PMID:27527519

  18. Human Frataxin Folds Via an Intermediate State. Role of the C-Terminal Region

    NASA Astrophysics Data System (ADS)

    Faraj, Santiago E.; González-Lebrero, Rodolfo M.; Roman, Ernesto A.; Santos, Javier

    2016-02-01

    The aim of this study is to investigate the folding reaction of human frataxin, whose deficiency causes the neurodegenerative disease Friedreich’s Ataxia (FRDA). The characterization of different conformational states would provide knowledge about how frataxin can be stabilized without altering its functionality. Wild-type human frataxin and a set of mutants, including two highly destabilized FRDA-associated variants were studied by urea-induced folding/unfolding in a rapid mixing device and followed by circular dichroism. The analysis clearly indicates the existence of an intermediate state (I) in the folding route with significant secondary structure content but relatively low compactness, compared with the native ensemble. However, at high NaCl concentrations I-state gains substantial compaction, and the unfolding barrier is strongly affected, revealing the importance of electrostatics in the folding mechanism. The role of the C-terminal region (CTR), the key determinant of frataxin stability, was also studied. Simulations consistently with experiments revealed that this stretch is essentially unstructured, in the most compact transition state ensemble (TSE2). The complete truncation of the CTR drastically destabilizes the native state without altering TSE2. Results presented here shed light on the folding mechanism of frataxin, opening the possibility of mutating it to generate hyperstable variants without altering their folding kinetics.

  19. Four residues of propeptide are essential for precursor folding of nattokinase.

    PubMed

    Jia, Yan; Cao, Xinhua; Deng, Yu; Bao, Wei; Tang, Changyan; Ding, Hanjing; Zheng, Zhongliang; Zou, Guolin

    2014-11-01

    Subtilisin propeptide functions as an intramolecular chaperone that guides precursor folding. Nattokinase, a member of subtilisin family, is synthesized as a precursor consisting of a signal peptide, a propeptide, and a subtilisin domain, and the mechanism of its folding remains to be understood. In this study, the essential residues of nattokinase propeptide which contribute to precursor folding were determined. Deletion analysis showed that the conserved regions in propeptide were important for precursor folding. Single-site and multi-site mutagenesis studies confirmed the role of Tyr10, Gly13, Gly34, and Gly35. During stage (i) and (ii) of precursor folding, Tyr10 and Gly13 would form the part of interface with subtilisin domain. While Gly34 and Gly35 connected with an α-helix that would stabilize the structure of propeptide. The quadruple Ala mutation, Y10A/G13A/G34A/G35A, resulted in a loss of the chaperone function for the propeptide. This work showed the essential residues of propeptide for precursor folding via secondary structure and kinetic parameter analyses.

  20. Human Frataxin Folds Via an Intermediate State. Role of the C-Terminal Region

    PubMed Central

    Faraj, Santiago E.; González-Lebrero, Rodolfo M.; Roman, Ernesto A.; Santos, Javier

    2016-01-01

    The aim of this study is to investigate the folding reaction of human frataxin, whose deficiency causes the neurodegenerative disease Friedreich’s Ataxia (FRDA). The characterization of different conformational states would provide knowledge about how frataxin can be stabilized without altering its functionality. Wild-type human frataxin and a set of mutants, including two highly destabilized FRDA-associated variants were studied by urea-induced folding/unfolding in a rapid mixing device and followed by circular dichroism. The analysis clearly indicates the existence of an intermediate state (I) in the folding route with significant secondary structure content but relatively low compactness, compared with the native ensemble. However, at high NaCl concentrations I-state gains substantial compaction, and the unfolding barrier is strongly affected, revealing the importance of electrostatics in the folding mechanism. The role of the C-terminal region (CTR), the key determinant of frataxin stability, was also studied. Simulations consistently with experiments revealed that this stretch is essentially unstructured, in the most compact transition state ensemble (TSE2). The complete truncation of the CTR drastically destabilizes the native state without altering TSE2. Results presented here shed light on the folding mechanism of frataxin, opening the possibility of mutating it to generate hyperstable variants without altering their folding kinetics. PMID:26856628

  1. Human telomeric G-quadruplex: thermodynamic and kinetic studies of telomeric quadruplex stability

    PubMed Central

    Chaires, Jonathan B.

    2010-01-01

    Summary Thermodynamic and kinetic studies complement high-resolution structures of G-quadruplexes. Such studies are essential for a thorough understanding of the mechanisms that govern quadruplex folding and conformational changes in quadruplexes. This perspective article reviews representative thermodynamic and kinetic studies of the folding of human telomeric quadruplex structures. Published thermodynamic data vary widely and are inconsistent. Possible reasons for these inconsistencies are discussed. The key issue of whether or not such folding reactions are a simple two-state process is examined. A tentative energy balance for the folding of telomeric quadruplexes in Na+ and K+ solution, and for conformational transition between these forms will be presented. PMID:19951355

  2. Efficient traversal of beta-sheet protein folding pathways using ensemble models.

    PubMed

    Shenker, Solomon; O'Donnell, Charles W; Devadas, Srinivas; Berger, Bonnie; Waldispühl, Jérôme

    2011-11-01

    Molecular dynamics (MD) simulations can now predict ms-timescale folding processes of small proteins; however, this presently requires hundreds of thousands of CPU hours and is primarily applicable to short peptides with few long-range interactions. Larger and slower-folding proteins, such as many with extended β-sheet structure, would require orders of magnitude more time and computing resources. Furthermore, when the objective is to determine only which folding events are necessary and limiting, atomistic detail MD simulations can prove unnecessary. Here, we introduce the program tFolder as an efficient method for modelling the folding process of large β-sheet proteins using sequence data alone. To do so, we extend existing ensemble β-sheet prediction techniques, which permitted only a fixed anti-parallel β-barrel shape, with a method that predicts arbitrary β-strand/β-strand orientations and strand-order permutations. By accounting for all partial and final structural states, we can then model the transition from random coil to native state as a Markov process, using a master equation to simulate population dynamics of folding over time. Thus, all putative folding pathways can be energetically scored, including which transitions present the greatest barriers. Since correct folding pathway prediction is likely determined by the accuracy of contact prediction, we demonstrate the accuracy of tFolder to be comparable with state-of-the-art methods designed specifically for the contact prediction problem alone. We validate our method for dynamics prediction by applying it to the folding pathway of the well-studied Protein G. With relatively very little computation time, tFolder is able to reveal critical features of the folding pathways which were only previously observed through time-consuming MD simulations and experimental studies. Such a result greatly expands the number of proteins whose folding pathways can be studied, while the algorithmic integration of

  3. Fold in Origami and Unfold Math

    ERIC Educational Resources Information Center

    Georgeson, Joseph

    2011-01-01

    Students enjoy origami and like making everything from paper cranes to footballs out of small, colorful squares of paper. They can invent their own shapes and are intrigued by the polyhedrons that they can construct. Paper folding is fun, but where is the math? Unless teachers develop lessons that address mathematical objectives, origami could be…

  4. Role of cofactors in metalloprotein folding.

    PubMed

    Wilson, Corey J; Apiyo, David; Wittung-Stafshede, Pernilla

    2004-01-01

    Metals are commonly found as natural constituents of proteins. Since many such metals can interact specifically with their corresponding unfolded proteins in vitro , cofactor-binding prior to polypeptide folding may be a biological path to active metalloproteins. By interacting with the unfolded polypeptide, the metal may create local structure that initiates and directs the polypeptide-folding process. Here, we review recent literature that addresses the involvement of metals in protein-folding reactions in vitro . To date, the best characterized systems are simple one such as blue-copper proteins, heme-binding proteins, iron-sulfur-cluster proteins and synthetic metallopeptides. Taken together, the available data demonstrates that metals can play diverse roles: it is clear that many cofactors bind before polypeptide folding and influence the reaction; yet, some do not bind until a well-structured active site is formed. The significance of characterizing the effects of metals on protein conformational changes is underscored by the many human diseases that are directly linked to anomalous protein-metal interactions.

  5. Folded-path optical analysis gas cell

    DOEpatents

    Carangelo, Robert M.; Wright, David D.

    1995-01-01

    A folded-path gas cell employs an elliptical concave mirror in confronting relationship to two substantially spherical concave mirrors. At least one of the spherical mirrors, and usually both, are formed with an added cylindrical component to increase orthogonal focii coincidence and thereby to increase the radiation energy throughput characteristic of the cell.

  6. Hydroxyapatite surface-induced peptide folding.

    PubMed

    Capriotti, Lisa A; Beebe, Thomas P; Schneider, Joel P

    2007-04-25

    Herein, we describe the design and surface-binding characterization of a de novo designed peptide, JAK1, which undergoes surface-induced folding at the hydroxyapatite (HA)-solution interface. JAK1 is designed to be unstructured in buffered saline solution, yet undergo HA-induced folding that is largely governed by the periodic positioning of gamma-carboxyglutamic acid (Gla) residues within the primary sequence of the peptide. Circular dichroism (CD) spectroscopy and analytical ultracentrifugation indicate that the peptide remains unfolded and monomeric in solution under normal physiological conditions; however, CD spectroscopy indicates that in the presence of hydroxyapatite, the peptide avidly binds to the mineral surface adopting a helical structure. Adsorption isotherms indicate nearly quantitative surface coverage and Kd = 310 nM for the peptide-surface binding event. X-ray photoelectron spectroscopy (XPS) coupled with the adsorption isotherm data suggests that JAK1 binds to HA, forming a self-limiting monolayer. This study demonstrates the feasibility of using HA surfaces to trigger the intramolecular folding of designed peptides and represents the initial stages of defining the design rules that allow HA-induced peptide folding.

  7. Folding and faulting of an elastic continuum

    PubMed Central

    Gourgiotis, Panos A.

    2016-01-01

    Folding is a process in which bending is localized at sharp edges separated by almost undeformed elements. This process is rarely encountered in Nature, although some exceptions can be found in unusual layered rock formations (called ‘chevrons’) and seashell patterns (for instance Lopha cristagalli). In mechanics, the bending of a three-dimensional elastic solid is common (for example, in bulk wave propagation), but folding is usually not achieved. In this article, the route leading to folding is shown for an elastic solid obeying the couple-stress theory with an extreme anisotropy. This result is obtained with a perturbation technique, which involves the derivation of new two-dimensional Green's functions for applied concentrated force and moment. While the former perturbation reveals folding, the latter shows that a material in an extreme anisotropic state is also prone to a faulting instability, in which a displacement step of finite size emerges. Another failure mechanism, namely the formation of dilation/compaction bands, is also highlighted. Finally, a geophysical application to the mechanics of chevron formation shows how the proposed approach may explain the formation of natural structures. PMID:27118925

  8. How do metal ions direct ribozyme folding?

    NASA Astrophysics Data System (ADS)

    Denesyuk, Natalia A.; Thirumalai, D.

    2015-10-01

    Ribozymes, which carry out phosphoryl-transfer reactions, often require Mg2+ ions for catalytic activity. The correct folding of the active site and ribozyme tertiary structure is also regulated by metal ions in a manner that is not fully understood. Here we employ coarse-grained molecular simulations to show that individual structural elements of the group I ribozyme from the bacterium Azoarcus form spontaneously in the unfolded ribozyme even at very low Mg2+ concentrations, and are transiently stabilized by the coordination of Mg2+ ions to specific nucleotides. However, competition for scarce Mg2+ and topological constraints that arise from chain connectivity prevent the complete folding of the ribozyme. A much higher Mg2+ concentration is required for complete folding of the ribozyme and stabilization of the active site. When Mg2+ is replaced by Ca2+ the ribozyme folds, but the active site remains unstable. Our results suggest that group I ribozymes utilize the same interactions with specific metal ligands for both structural stability and chemical activity.

  9. Hydroxyapatite surface-induced peptide folding.

    PubMed

    Capriotti, Lisa A; Beebe, Thomas P; Schneider, Joel P

    2007-04-25

    Herein, we describe the design and surface-binding characterization of a de novo designed peptide, JAK1, which undergoes surface-induced folding at the hydroxyapatite (HA)-solution interface. JAK1 is designed to be unstructured in buffered saline solution, yet undergo HA-induced folding that is largely governed by the periodic positioning of gamma-carboxyglutamic acid (Gla) residues within the primary sequence of the peptide. Circular dichroism (CD) spectroscopy and analytical ultracentrifugation indicate that the peptide remains unfolded and monomeric in solution under normal physiological conditions; however, CD spectroscopy indicates that in the presence of hydroxyapatite, the peptide avidly binds to the mineral surface adopting a helical structure. Adsorption isotherms indicate nearly quantitative surface coverage and Kd = 310 nM for the peptide-surface binding event. X-ray photoelectron spectroscopy (XPS) coupled with the adsorption isotherm data suggests that JAK1 binds to HA, forming a self-limiting monolayer. This study demonstrates the feasibility of using HA surfaces to trigger the intramolecular folding of designed peptides and represents the initial stages of defining the design rules that allow HA-induced peptide folding. PMID:17397165

  10. Folded-path optical analysis gas cell

    DOEpatents

    Carangelo, R.M.; Wright, D.D.

    1995-08-08

    A folded-path gas cell employs an elliptical concave mirror in confronting relationship to two substantially spherical concave mirrors. At least one of the spherical mirrors, and usually both, are formed with an added cylindrical component to increase orthogonal foci coincidence and thereby to increase the radiation energy throughput characteristic of the cell. 10 figs.

  11. Folded cavity design for a ruby resonator

    NASA Technical Reports Server (NTRS)

    Arunkumar, K. A.; Trolinger, James D.

    1988-01-01

    A folded cavity laser resonator operating in the TEM(00) mode has been built and tested. The new oscillator configuration leads to an increase in efficiency and to better line narrowing due to the increased number of passes through the laser rod and tuning elements, respectively. The modification is shown to lead to cavity ruggedization.

  12. Fast phase randomization via two-folds

    PubMed Central

    Jeffrey, M. R.

    2016-01-01

    A two-fold is a singular point on the discontinuity surface of a piecewise-smooth vector field, at which the vector field is tangent to the discontinuity surface on both sides. If an orbit passes through an invisible two-fold (also known as a Teixeira singularity) before settling to regular periodic motion, then the phase of that motion cannot be determined from initial conditions, and, in the presence of small noise, the asymptotic phase of a large number of sample solutions is highly random. In this paper, we show how the probability distribution of the asymptotic phase depends on the global nonlinear dynamics. We also show how the phase of a smooth oscillator can be randomized by applying a simple discontinuous control law that generates an invisible two-fold. We propose that such a control law can be used to desynchronize a collection of oscillators, and that this manner of phase randomization is fast compared with existing methods (which use fixed points as phase singularities), because there is no slowing of the dynamics near a two-fold. PMID:27118901

  13. Global hairpin folding of tau in solution.

    PubMed

    Jeganathan, Sadasivam; von Bergen, Martin; Brutlach, Henrik; Steinhoff, Heinz-Jürgen; Mandelkow, Eckhard

    2006-02-21

    The microtubule-associated protein tau stabilizes microtubules in its physiological role, whereas it forms insoluble aggregates (paired helical filaments) in Alzheimer's disease. Soluble tau is considered a natively unfolded protein whose residual folding and intramolecular interactions are largely undetermined. In this study, we have applied fluorescence resonance energy transfer (FRET) and electron paramagnetic resonance (EPR) to examine the proximity and flexibility of tau domains and the global folding. FRET pairs spanning the tau molecule were created by inserting tryptophans (donor) and cysteines (labeled with IAEDANS as an acceptor) by site-directed mutagenesis. The observed FRET distances were significantly different from those expected for a random coil. Notably, the C-terminal end of tau folds over into the vicinity of the microtubule-binding repeat domain, the N-terminus remains outside the FRET distance of the repeat domain, yet both ends of the molecule approach one another. The interactions between the domains were obliterated by denaturation in GdnHCl. Paramagnetic spin-labels attached in various domains of tau were analyzed by EPR and exhibited a high mobility throughout. The data indicate that tau retains some global folding even in its "natively unfolded" state, combined with the high flexibility of the chain.

  14. Self-folding graphene-polymer bilayers

    NASA Astrophysics Data System (ADS)

    Deng, Tao; Yoon, ChangKyu; Jin, Qianru; Li, Mingen; Liu, Zewen; Gracias, David H.

    2015-05-01

    In order to incorporate the extraordinary intrinsic thermal, electrical, mechanical, and optical properties of graphene with three dimensional (3D) flexible substrates, we introduce a solvent-driven self-folding approach using graphene-polymer bilayers. A polymer (SU-8) film was spin coated atop chemically vapor deposited graphene films on wafer substrates and graphene-polymer bilayers were patterned with or without metal electrodes using photolithography, thin film deposition, and etching. After patterning, the bilayers were released from the substrates and they self-folded to form fully integrated, curved, and folded structures. In contrast to planar graphene sensors on rigid substrates, we assembled curved and folded sensors that are flexible and they feature smaller form factors due to their 3D geometry and large surface areas due to their multiple rolled architectures. We believe that this approach could be used to assemble a range of high performance 3D electronic and optical devices of relevance to sensing, diagnostics, wearables, and energy harvesting.

  15. Self-folding graphene-polymer bilayers

    SciTech Connect

    Deng, Tao; Yoon, ChangKyu; Jin, Qianru; Li, Mingen; Liu, Zewen; Gracias, David H.

    2015-05-18

    In order to incorporate the extraordinary intrinsic thermal, electrical, mechanical, and optical properties of graphene with three dimensional (3D) flexible substrates, we introduce a solvent-driven self-folding approach using graphene-polymer bilayers. A polymer (SU-8) film was spin coated atop chemically vapor deposited graphene films on wafer substrates and graphene-polymer bilayers were patterned with or without metal electrodes using photolithography, thin film deposition, and etching. After patterning, the bilayers were released from the substrates and they self-folded to form fully integrated, curved, and folded structures. In contrast to planar graphene sensors on rigid substrates, we assembled curved and folded sensors that are flexible and they feature smaller form factors due to their 3D geometry and large surface areas due to their multiple rolled architectures. We believe that this approach could be used to assemble a range of high performance 3D electronic and optical devices of relevance to sensing, diagnostics, wearables, and energy harvesting.

  16. Protein fold recognition using geometric kernel data fusion

    PubMed Central

    Zakeri, Pooya; Jeuris, Ben; Vandebril, Raf; Moreau, Yves

    2014-01-01

    Motivation: Various approaches based on features extracted from protein sequences and often machine learning methods have been used in the prediction of protein folds. Finding an efficient technique for integrating these different protein features has received increasing attention. In particular, kernel methods are an interesting class of techniques for integrating heterogeneous data. Various methods have been proposed to fuse multiple kernels. Most techniques for multiple kernel learning focus on learning a convex linear combination of base kernels. In addition to the limitation of linear combinations, working with such approaches could cause a loss of potentially useful information. Results: We design several techniques to combine kernel matrices by taking more involved, geometry inspired means of these matrices instead of convex linear combinations. We consider various sequence-based protein features including information extracted directly from position-specific scoring matrices and local sequence alignment. We evaluate our methods for classification on the SCOP PDB-40D benchmark dataset for protein fold recognition. The best overall accuracy on the protein fold recognition test set obtained by our methods is ∼86.7%. This is an improvement over the results of the best existing approach. Moreover, our computational model has been developed by incorporating the functional domain composition of proteins through a hybridization model. It is observed that by using our proposed hybridization model, the protein fold recognition accuracy is further improved to 89.30%. Furthermore, we investigate the performance of our approach on the protein remote homology detection problem by fusing multiple string kernels. Availability and implementation: The MATLAB code used for our proposed geometric kernel fusion frameworks are publicly available at http://people.cs.kuleuven.be/∼raf.vandebril/homepage/software/geomean.php?menu=5/ Contact: pooyapaydar@gmail.com or yves

  17. Description and classification of folds in single surfaces

    NASA Astrophysics Data System (ADS)

    Twiss, Robert J.

    I propose a new three-parameter description of fold style in folded surfaces based on the ratio of the amplitude to the half-wavelength (the aspect ratio P), the maximum angle of relative rotation of opposite limbs of the fold (the folding angle φ), and a measure of the relative curvature at the fold closure (the bluntness b). For symmetric folds, the first two parameters define a trapezoid that circumscribes the fold and provides the primary criterion for the classification of fold style. Within a given trapezoid, fold style variations are defined by the bluntness. Perfect folds in profile are defined to have a single hinge with perfectly straight limbs tangent to hinge zones that are perfect circular arcs. An analytic description of the variation in perfect fold geometry defines the limits for all natural single-hinged folds. The proposed system includes folds with folding angles both less than and greater than isoclinal folds, it applies to both single-hinged and multiple-hinged folds, and it also can be extended to apply to asymmetric folds. Previously proposed two-parameter classification systems can only describe folds that are restricted to a specific surface through the three-parameter fold style space proposed here.

  18. Fault-related folding during extension: Plunging basement-cored folds in the Basin and Range

    USGS Publications Warehouse

    Howard, K.A.; John, Barbara E.

    1997-01-01

    Folds are able to form in highly extended areas where stratified cover rocks respond to basement fault offsets. The response of cover rocks to basement faulting can be studied especially well in plunging structures that expose large structural relief. The southern Basin and Range province contains plunging folds kilometres in amplitude at the corners of domino-like tilt blocks of basement rocks, where initially steep transverse and normal faults propagated upward toward the layered cover rocks. Exposed tilted cross sections, as much as 8 km thick, display transitions from faulted basement to folded cover that validate laboratory models of forced folds. The folded cover masks a deeper extensional style of brittle segmentation and uniform steep tilting.

  19. Evolutionary conservation of codon optimality reveals hidden signatures of cotranslational folding.

    PubMed

    Pechmann, Sebastian; Frydman, Judith

    2013-02-01

    The choice of codons can influence local translation kinetics during protein synthesis. Whether codon preference is linked to cotranslational regulation of polypeptide folding remains unclear. Here, we derive a revised translational efficiency scale that incorporates the competition between tRNA supply and demand. Applying this scale to ten closely related yeast species, we uncover the evolutionary conservation of codon optimality in eukaryotes. This analysis reveals universal patterns of conserved optimal and nonoptimal codons, often in clusters, which associate with the secondary structure of the translated polypeptides independent of the levels of expression. Our analysis suggests an evolved function for codon optimality in regulating the rhythm of elongation to facilitate cotranslational polypeptide folding, beyond its previously proposed role of adapting to the cost of expression. These findings establish how mRNA sequences are generally under selection to optimize the cotranslational folding of corresponding polypeptides.

  20. Architecture and Folding Mechanism of the Azoarcus Group I Pre-tRNA

    SciTech Connect

    Rangan,P.; Masquida, B.; Westhof, E.; Woodson, S.

    2004-01-01

    Self-splicing RNAs must evolve to function in their specific exon context. The conformation of a group I pre-tRNA{sup ile} from the bacterium Azoarcus was probed by ribonuclease T1 and hydroxyl radical cleavage, and by native gel electrophoresis. Biochemical data and three-dimensional models of the pre-tRNA showed that the tRNA is folded, and that the tRNA and intron sequences form separate tertiary domains. Models of the active site before steps 1 and 2 of the splicing reaction predict that exchange of the external G-cofactor and the 3{prime}-terminal G is accomplished by a slight conformational change in P9.0 of the Azoarcus group I intron. Kinetic assays showed that the pre-tRNA folds in minutes, much more slowly than the intron alone. The dependence of the folding kinetics on Mg{sup 2+} and the concentration of urea, and RNase T1 experiments showed that formation of native pre-tRNA is delayed by misfolding of P3-P9, including mispairing between residues in P9 and the tRNA. Thus, although the intron and tRNA sequences form separate domains in the native pre-tRNA, their folding is coupled via metastable non-native base-pairs. This could help prevent premature processing of the 5{prime} and 3{prime} ends of unspliced pre-tRNA.

  1. Formation of Tertiary Interactions during rRNA GTPase Center Folding.

    PubMed

    Rau, Michael J; Welty, Robb; Tom Stump, W; Hall, Kathleen B

    2015-08-28

    The 60-nt 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 Mg(2+) ions, which is stabilized by the L11 protein in cocrystal structures. Here, we monitor the kinetics of its tertiary folding and Mg(2+)-dependent intermediate states by observing selected nucleobases that contribute specific interactions to the GAC tertiary structure in the cocrystals. The fluorescent nucleobase 2-aminopurine 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 Mg(2+) 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 to 4s and with global folding from 1 to 5s. Site-specific substitution with (15)N-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 Mg(2+) and suggest that there are two initial sites of Mg(2+) ion association. PMID:26210661

  2. Thermodynamics and Kinetics of RNA Unfolding and Refolding

    NASA Astrophysics Data System (ADS)

    Li, Pan T. X.; Tinoco, Ignacio

    Emergence of novel functions of non-coding RNAs imposes a new challenge for thermodynamics-based structural prediction of RNA. Here we review bulk and single-molecule techniques to measure the thermodynamics and kinetics of RNA folding and unfolding. RNA can be denatured by heat, chemicals, force, and by depletion of divalent cations. Various spectroscopic, calorimetric, chemical and biochemical methods have been used to study RNA structures. We emphasize single-molecule force unfolding as a new and powerful technique to study RNA structure and folding. Using optical tweezers, single RNA molecules can be stretched and relaxed; their changes in extension reflect structural rearrangements. We discuss determination of Gibb free energy of folding from mechanical work under both equilibrium and non-equilibrium conditions. Force can be applied to affect reaction rates as well as to manipulate molecular structure. Folding and unfolding kinetics can be monitored in real time.

  3. Geometric models of folding at Loch Monar, Scotland, using computer simulation

    NASA Astrophysics Data System (ADS)

    Watkinson, A. J.; Thiessen, R. L.

    1988-06-01

    Small-scale hand specimens were collected at Loch Monar, Scotland, one of the classic areas of fold interference patterns. From the analysis of these, the geometries of the F 1 and F 2 folds were derived. By computer simulation, the complex three-dimensional form of the fold interference shapes can then be reproduced very closely. F 2 fold shapes and motion directions derived from the small-scale structures, along with those derived from field observations of F 1 lineations deformed about F 2 folds, were then applied to an extrapolated pre-F 2 shape of the Loch Monar synform. This generated a map pattern strikingly similar to that mapped by Ramsay (1958). The geometric models provide useful information for mechanical hypotheses of the folding observed at Loch Monar. The simulation reveals an interesting problem that whereas many small-scale interference patterns reflect the map pattern of the major structure, there are exposures of interference patterns that do not. These are dome and basin (type 1) patterns found in an area where a major F 2 fold hinge crosses an F 1 fold hinge zone. By examining the deformed L 1 lineation patterns found in this area, along with the computed D 2 strain orientations and the field observations of F 2 fold geometries, we suggest that those patterns formed due to local variations in the displacement directions during the F 2 folding, perhaps due to the mechanical influence of relatively competent pegmatite veins on the small-scale F 2 folds. The very high D 2 strain has then amplified the dome and basin elements to very elongate cone and cylinder forms. This creates local type 1 patterns within a regional type 2 interference pattern structure.

  4. Generation of buckle folds in Naga fold thrust belt, north-east India

    NASA Astrophysics Data System (ADS)

    Saha, B.; Dietl, C.

    2009-04-01

    Naga fold thrust belt (NFTB), India, formed as a result of northward migration of the Indian plate initiated in Eocene and its subsequent collision with the Burmese plate during Oligocene. The NW-SE oriented compression generated a spectrum of structures; among them, we intend to focus on the folds- varying from gentle to tight asymmetric in geometry. Large recumbent folds are often associated with thrusting. Buckle folds forming under shallow crustal conditions are frequently reported from NFTB. Buckle folding occurs mainly within sandstones with intercalated shale layers which are in the study area typical for the Barail, Surma and Tipam Groups. We have tried to explain the controlling factors behind the variation of the buckle fold shapes and their varying wavelengths throughout the fold thrust belt with the aid of analogue (sand box) modelling. It is undoubted that competence contrast along with the layer parallel compressive stress are the major influencing factors in generation of buckle folds. Schmalholz and Podladchikov (1999) and Jeng et al. (2002) have shown that when low strain rate and low temperature are applicable, not only the viscosity contrast, but also the elasticity contrast govern the geometry of the developing buckle folds. Rocks deforming under high temperature and high pressure deform in pure viscous manner, whereas, rocks undergoing less confining stress and less temperature, are subjected to pure elastic deformation. However, they are the end members, and most of the deformations are a combination of these two end members, i.e. of viscoelastic nature. Our models are made up of sieved sand (0.5 mm grain size) and mica layers (1-5 mm) This interlayering imparts a mechanical anisotropy in the model. Mica is not a pure viscous material, rather it displays more elastic behaviour. The mica layers in the model produce bedding parallel slip during shortening through internal reorganization of the individual mica crystals leading to the thickening

  5. Local rules for protein folding on a triangular lattice and generalized hydrophobicity in the HP model

    SciTech Connect

    Agarwala, R.; Batzoglou, S.; Dancik, V.

    1997-06-01

    We consider the problem of determining the three-dimensional folding of a protein given its one-dimensional amino acid sequence. We use the HP model for protein folding proposed by Dill, which models protein as a chain of amino acid residues that are either hydrophobic or polar, and hydrophobic interactions are the dominant initial driving force for the protein folding. Hart and Istrail gave approximation algorithms for folding proteins on the cubic lattice under HP model. In this paper, we examine the choice of a lattice by considering its algorithmic and geometric implications and argue that triangular lattice is a more reasonable choice. We present a set of folding rules for a triangular lattice and analyze the approximation ratio which they achieve. In addition, we introduce a generalization of the HP model to account for residues having different levels of hydrophobicity. After describing the biological foundation for this generalization, we show that in the new model we are able to achieve similar constant factor approximation guarantees on the triangular lattice as were achieved in the standard HP model. While the structures derived from our folding rules are probably still far from biological reality, we hope that having a set of folding rules with different properties will yield more interesting folds when combined.

  6. Laboratory evolution of fast-folding green fluorescent protein using secretory pathway quality control.

    PubMed

    Fisher, Adam C; DeLisa, Matthew P

    2008-01-01

    Green fluorescent protein (GFP) has undergone a long history of optimization to become one of the most popular proteins in all of cell biology. It is thermally and chemically robust and produces a pronounced fluorescent phenotype when expressed in cells of all types. Recently, a superfolder GFP was engineered with increased resistance to denaturation and improved folding kinetics. Here we report that unlike other well-folded variants of GFP (e.g., GFPmut2), superfolder GFP was spared from elimination when targeted for secretion via the SecYEG translocase. This prompted us to hypothesize that the folding quality control inherent to this secretory pathway could be used as a platform for engineering similar 'superfolded' proteins. To test this, we targeted a combinatorial library of GFPmut2 variants to the SecYEG translocase and isolated several superfolded variants that accumulated in the cytoplasm due to their enhanced folding properties. Each of these GFP variants exhibited much faster folding kinetics than the parental GFPmut2 protein and one of these, designated superfast GFP, folded at a rate that even exceeded superfolder GFP. Remarkably, these GFP variants exhibited little to no loss in specific fluorescence activity relative to GFPmut2, suggesting that the process of superfolding can be accomplished without altering the proteins' normal function. Overall, we demonstrate that laboratory evolution combined with secretory pathway quality control enables sampling of largely unexplored amino-acid sequences for the discovery of artificial, high-performance proteins with properties that are unparalleled in their naturally occurring analogues. PMID:18545653

  7. Reversible Aggregation Plays a Crucial Role on the Folding Landscape of p53 Core Domain

    PubMed Central

    Ishimaru, Daniella; Lima, Luis M. T. R.; Maia, Lenize F.; Lopez, Priscila M.; Ano Bom, Ana P.; Valente, Ana P.; Silva, Jerson L.

    2004-01-01

    The role of tumor suppressor protein p53 in cell cycle control depends on its flexible and partially unstructured conformation, which makes it crucial to understand its folding landscape. Here we report an intermediate structure of the core domain of the tumor suppressor protein p53 (p53C) during equilibrium and kinetic folding/unfolding transitions induced by guanidinium chloride. This partially folded structure was undetectable when investigated by intrinsic fluorescence. Indeed, the fluorescence data showed a simple two-state transition. On the other hand, analysis of far ultraviolet circular dichroism in 1.0 M guanidinium chloride demonstrated a high content of secondary structure, and the use of an extrinsic fluorescent probe, 4,4′-dianilino-1,1′ binaphthyl-5,5′-disulfonic acid, indicated an increase in exposure of the hydrophobic core at 1 M guanidinium chloride. This partially folded conformation of p53C was plagued by aggregation, as suggested by one-dimensional NMR and demonstrated by light-scattering and gel-filtration chromatography. Dissociation by high pressure of these aggregates reveals the reversibility of the process and that the aggregates have water-excluded cavities. Kinetic measurements show that the intermediate formed in a parallel reaction between unfolded and folded structures and that it is under fine energetic control. They are not only crucial to the folding pathway of p53C but may explain as well the vulnerability of p53C to undergo departure of the native to an inactive state, which makes the cell susceptible to malignant transformation. PMID:15298872

  8. One-Dimensional Barrier-Preserving Free-Energy Projections of β-sheet Miniprotein: New Insights into the Folding Process

    PubMed Central

    Krivov, Sergei V.; Muff, Stefanie; Caflisch, Amedeo; Karplus, Martin

    2009-01-01

    The conformational space of a 20-residue three-stranded antiparallel β-sheet peptide (double hairpin) was sampled by equilibrium folding/unfolding molecular dynamics simulations for a total of 20 µs. The resulting one-dimensional free-energy profiles (FEPs) provide a detailed description of the free-energy basins and barriers for the folding reaction. The similarity of the FEPs obtained using the probability of folding before unfolding (p fold) or the mean first passage time supports the robustness of the procedure. The folded state and the most populated free-energy basins in the denatured state are described by the one-dimensional FEPs, which avoid the overlap of states present in the usual one- or two-dimensional projections. Within the denatured state, a basin with fluctuating helical conformations and a heterogeneous entropic state are populated near the melting temperature at about 11% and 33%, respectively. Folding pathways from the helical basin or enthalpic traps (with only one of the two hairpins formed) reach the native state through the entropic state, which is on-pathway and is separated by a low barrier from the folded state. A simplified equilibrium kinetic network based on the FEPs shows the complexity of the folding reaction and indicates, as augmented by additional analyses, that the basins in the denatured state are connected primarily by the native state. The overall folding kinetics shows single-exponential behavior because barriers between the non-native basins and the folded state have similar heights. PMID:18590307

  9. Oxidation of kinetically trapped thiols by protein disulfide isomerase.

    PubMed

    Walker, K W; Gilbert, H F

    1995-10-17

    The formation of a stabilized structure during oxidative protein folding can severely retard disulfide formation if the structure must be disrupted to gain access to buried cysteines. These kinetic traps can slow protein folding and disulfide bond formation to the extent that unassisted folding is too slow to be kinetically competent in the cell. Protein disulfide isomerase (PDI) facilitates the oxidation of a kinetically trapped state of RTEM-1 beta-lactamase in which two cysteines that form the single disulfide bond in the native protein are buried and approximately 500-fold less reactive than exposed cysteines. Under second-order conditions, PDI-dependent oxidation of reduced, folded beta-lactamase is 500-fold faster than GSSG-dependent oxidation. The rate difference observed between PDI and GSSG can be accounted for by the 520-fold higher kinetic reactivity of PDI as an oxidant. Noncovalent interactions between PDI (35 microM) and beta-lactamase increase the reactivity or unfolding of beta-lactamase in the steady-state by less than 3-fold. At high concentrations of PDI or alkylating agents, the reaction of beta-lactamase cysteines approaches a constant rate, limited by the spontaneous unfolding of the protein (kunfold = 0.024 +/- 0.005 min-1). PDI does not substantially increase the rate of beta-lactamase unfolding; however, once beta-lactamase spontaneously unfolds, PDI at concentrations greater than 44 +/- 4 microM, oxidizes the unfolded substrate before it can refold (kfold = 1.5 +/- 0.2 min-1).(ABSTRACT TRUNCATED AT 250 WORDS)

  10. Unfolding a Problem

    ERIC Educational Resources Information Center

    Currier, Sarah Cox

    2015-01-01

    In this article, Sarah Currier, a math specialist at Elizabeth Hall International School in Minnesota, describes how she used origami in a deliberate manner to teach content. She shares how she uses paper folding to teach mathematical concepts, reinforce vocabulary, and as a problem-solving model. She also offers ideas for using origami in other…

  11. Topological Constraints on Chain-Folding Structure of Semicrystalline Polymer as Studied by 13C-13C Double Quantum NMR

    NASA Astrophysics Data System (ADS)

    Hong, Youlee; Miyoshi, Toshikazu

    Chain-folding process is a prominent feature of long polymer chains during crystallization. Over the last half century, much effort has been paid to reveal the chain trajectory. Even though various chain-folding models as well as theories of crystallization at molecule levels have been proposed, they could be not reconciled due to the limited experimental evidences. Recent development of double quantum NMR with selective isotope labeling identified the chain-trajectory of 13C labeled isotactic poly(1-butene). The systematic experiments covered a wide range of parameters, i.e. kinetics, concentration, and molecular weight (Mw) . It was demonstrated that i) adjacent re-entry site was invariant as a function of crystallization temperature (Tc) , concentration, andMw, ii) long-range order of adjacent re-entry sequence is independence of kinetics at a given concentration while it decreased with increasing the polymer concentration at a given Tc due to the increased interruption between the chains, and iii) high Mw chains led to the multilayer folded structures in single crystals, but the melt state induced the identical short adjacent sequences of long and short polymer over a wide range of Tc due to the entanglements. The behaviors indicated that the topological restriction plays significant roles in the chain-folding process rather than the kinetics. The proposed framework to control the chain-folding structure presents a new perspective into the chain organization by either the intra- or inter-chain interaction. National Science Foundation Grants DMR-1105829 and 1408855.

  12. Microsecond Rearrangements of Hydrophobic Clusters in an Initially Collapsed Globule Prime Structure Formation during the Folding of a Small Protein.

    PubMed

    Goluguri, Rama Reddy; Udgaonkar, Jayant B

    2016-07-31

    Determining how polypeptide chain collapse initiates structure formation during protein folding is a long standing goal. It has been challenging to characterize experimentally the dynamics of the polypeptide chain, which lead to the formation of a compact kinetic molten globule (MG) in about a millisecond. In this study, the sub-millisecond events that occur early during the folding of monellin from the guanidine hydrochloride-unfolded state have been characterized using multiple fluorescence and fluorescence resonance energy transfer probes. The kinetic MG is shown to form in a noncooperative manner from the unfolded (U) state as a result of at least three different processes happening during the first millisecond of folding. Initial chain compaction completes within the first 37μs, and further compaction occurs only after structure formation commences at a few milliseconds of folding. The transient nonnative and native-like hydrophobic clusters with side chains of certain residues buried form during the initial chain collapse and the nonnative clusters quickly disassemble. Subsequently, partial chain desolvation occurs, leading to the formation of a kinetic MG. The initial chain compaction and subsequent chain rearrangement appear to be barrierless processes. The two structural rearrangements within the collapsed globule appear to prime the protein for the actual folding transition. PMID:27370109

  13. Start2Fold: a database of hydrogen/deuterium exchange data on protein folding and stability

    PubMed Central

    Pancsa, Rita; Varadi, Mihaly; Tompa, Peter; Vranken, Wim F.

    2016-01-01

    Proteins fulfil a wide range of tasks in cells; understanding how they fold into complex three-dimensional (3D) structures and how these structures remain stable while retaining sufficient dynamics for functionality is essential for the interpretation of overall protein behaviour. Since the 1950's, solvent exchange-based methods have been the most powerful experimental means to obtain information on the folding and stability of proteins. Considerable expertise and care were required to obtain the resulting datasets, which, despite their importance and intrinsic value, have never been collected, curated and classified. Start2Fold is an openly accessible database (http://start2fold.eu) of carefully curated hydrogen/deuterium exchange (HDX) data extracted from the literature that is open for new submissions from the community. The database entries contain (i) information on the proteins investigated and the underlying experimental procedures and (ii) the classification of the residues based on their exchange protection levels, also allowing for the instant visualization of the relevant residue groups on the 3D structures of the corresponding proteins. By providing a clear hierarchical framework for the easy sharing, comparison and (re-)interpretation of HDX data, Start2Fold intends to promote a better understanding of how the protein sequence encodes folding and structure as well as the development of new computational methods predicting protein folding and stability. PMID:26582925

  14. Vocal fold and ventricular fold vibration in period-doubling phonation: physiological description and aerodynamic modeling.

    PubMed

    Bailly, Lucie; Henrich, Nathalie; Pelorson, Xavier

    2010-05-01

    Occurrences of period-doubling are found in human phonation, in particular for pathological and some singing phonations such as Sardinian A Tenore Bassu vocal performance. The combined vibration of the vocal folds and the ventricular folds has been observed during the production of such low pitch bass-type sound. The present study aims to characterize the physiological correlates of this acoustical production and to provide a better understanding of the physical interaction between ventricular fold vibration and vocal fold self-sustained oscillation. The vibratory properties of the vocal folds and the ventricular folds during phonation produced by a professional singer are analyzed by means of acoustical and electroglottographic signals and by synchronized glottal images obtained by high-speed cinematography. The periodic variation in glottal cycle duration and the effect of ventricular fold closing on glottal closing time are demonstrated. Using the detected glottal and ventricular areas, the aerodynamic behavior of the laryngeal system is simulated using a simplified physical modeling previously validated in vitro using a larynx replica. An estimate of the ventricular aperture extracted from the in vivo data allows a theoretical prediction of the glottal aperture. The in vivo measurements of the glottal aperture are then compared to the simulated estimations. PMID:21117769

  15. Discretized torsional dynamics and the folding of an RNA chain

    NASA Astrophysics Data System (ADS)

    Fernández, Ariel; Salthú, Rodolfo; Cendra, Hernán

    1999-08-01

    The aim of this work is to implement a discrete coarse codification of local torsional states of the RNA chain backbone in order to explore the long-time limit dynamics and ultimately obtain a coarse solution to the RNA folding problem. A discrete representation of the soft-mode dynamics is turned into an algorithm for a rough structure prediction. The algorithm itself is inherently parallel, as it evaluates concurrent folding possibilities by pattern recognition, but it may be implemented in a personal computer as a chain of perturbation-translation-renormalization cycles performed on a binary matrix of local topological constraints. This requires suitable representational tools and a periodic quenching of the dynamics for system renormalization. A binary coding of local topological constraints associated with each structural motif is introduced, with each local topological constraint corresponding to a local torsional state. This treatment enables us to adopt a computation time step far larger than hydrodynamic drag time scales. Accordingly, the solvent is no longer treated as a hydrodynamic drag medium. Instead we incorporate its capacity for forming local conformation-dependent dielectric domains. Each translation of the matrix of local topological constraints (LTM's) depends on the conformation-dependent local dielectric created by a confined solvent. Folding pathways are resolved as transitions between patterns of locally encoded structural signals which change within the 1 ns-100 ms time scale range. These coarse folding pathways are generated by a search at regular intervals for structural patterns in the LTM. Each pattern is recorded as a base-pairing pattern (BPP) matrix, a consensus-evaluation operation subject to a renormalization feedback loop. Since several mutually conflicting consensus evaluations might occur at a given time, the need arises for a probabilistic approach appropriate for an ensemble of RNA molecules. Thus, a statistical dynamics of

  16. The solution of pharmacological problems with computers. Part 9: Kinetics of the inhibition of the enzyme morphine-N-demethylase by N-allylnormorphine. Mathematical analysis by a FORTRAN-program.

    PubMed

    Krüger-Thiemer, E; Feinberg, M

    1975-03-01

    In this paper, we present a method for statistical analysis of enzyme-kinetic data. This analysis allows determination of the character of the inhibition of an enzyme reaction, and computes the 95% confidence limits of the dissociation constants and of Vmax. This method involves the solution of the Michaelis-Menten equation, (see article for formula), using the Guass-Newton iteration.

  17. Structure, stability and folding of the alpha-helix.

    PubMed

    Doig, A J; Andrew, C D; Cochran, D A; Hughes, E; Penel, S; Sun, J K; Stapley, B J; Clarke, D T; Jones, G R

    2001-01-01

    Pauling first described the alpha-helix nearly 50 years ago, yet new features of its structure continue to be discovered, using peptide model systems, site-directed mutagenesis, advances in theory, the expansion of the Protein Data Bank and new experimental techniques. Helical peptides in solution form a vast number of structures, including fully helical, fully coiled and partly helical. To interpret peptide results quantitatively it is essential to use a helix/coil model that includes the stabilities of all these conformations. Our models now include terms for helix interiors, capping, side-chain interactions, N-termini and 3(10)-helices. The first three amino acids in a helix (N1, N2 and N3) and the preceding N-cap are unique, as their amide NH groups do not participate in backbone hydrogen bonding. We surveyed their structures in proteins and measured their amino acid preferences. The results are predominantly rationalized by hydrogen bonding to the free NH groups. Stabilizing side-chain-side-chain energies, including hydrophobic interactions, hydrogen bonding and polar/non-polar interactions, were measured accurately in helical peptides. Helices in proteins show a preference for having approximately an integral number of turns so that their N- and C-caps lie on the same side. There are also strong periodic trends in the likelihood of terminating a helix with a Schellman or alpha L C-cap motif. The kinetics of alpha-helix folding have been studied with stopped-flow deep ultraviolet circular dichroism using synchrotron radiation as the light source; this gives a far superior signal-to-noise ratio than a conventional instrument. We find that poly(Glu), poly(Lys) and alanine-based peptides fold in milliseconds, with longer peptides showing a transient overshoot in helix content.

  18. Thermodynamic characterization of an equilibrium folding intermediate of staphylococcal nuclease.

    PubMed Central

    Xie, D.; Fox, R.; Freire, E.

    1994-01-01

    High-sensitivity differential scanning calorimetry and CD spectroscopy have been used to probe the structural stability and measure the folding/unfolding thermodynamics of a Pro117-->Gly variant of staphylococcal nuclease. It is shown that at neutral pH the thermal denaturation of this protein is well accounted for by a 2-state mechanism and that the thermally denatured state is a fully hydrated unfolded polypeptide. At pH 3.5, thermal denaturation results in a compact denatured state in which most, if not all, of the helical structure is missing and the beta subdomain apparently remains largely intact. At pH 3.0, no thermal transition is observed and the molecule exists in the compact denatured state within the 0-100 degrees C temperature interval. At high salt concentration and pH 3.5, the thermal unfolding transition exhibits 2 cooperative peaks in the heat capacity function, the first one corresponding to the transition from the native to the intermediate state and the second one to the transition from the intermediate to the unfolded state. As is the case with other proteins, the enthalpy of the intermediate is higher than that of the unfolded state at low temperatures, indicating that, under those conditions, its stabilization must be of an entropic origin. The folding intermediate has been modeled by structural thermodynamic calculations. Structure-based thermodynamic calculations also predict that the most probable intermediate is one in which the beta subdomain is essentially intact and the rest of the molecule unfolded, in agreement with the experimental data. The structural features of the equilibrium intermediate are similar to those of a kinetic intermediate previously characterized by hydrogen exchange and NMR spectroscopy. PMID:7756977

  19. Misplaced helix slows down ultrafast pressure-jump protein folding.

    PubMed

    Prigozhin, Maxim B; Liu, Yanxin; Wirth, Anna Jean; Kapoor, Shobhna; Winter, Roland; Schulten, Klaus; Gruebele, Martin

    2013-05-14

    Using a newly developed microsecond pressure-jump apparatus, we monitor the refolding kinetics of the helix-stabilized five-helix bundle protein λ*YA, the Y22W/Q33Y/G46,48A mutant of λ-repressor fragment 6-85, from 3 μs to 5 ms after a 1,200-bar P-drop. In addition to a microsecond phase, we observe a slower 1.4-ms phase during refolding to the native state. Unlike temperature denaturation, pressure denaturation produces a highly reversible helix-coil-rich state. This difference highlights the importance of the denatured initial condition in folding experiments and leads us to assign a compact nonnative helical trap as the reason for slower P-jump-induced refolding. To complement the experiments, we performed over 50 μs of all-atom molecular dynamics P-drop refolding simulations with four different force fields. Two of the force fields yield compact nonnative states with misplaced α-helix content within a few microseconds of the P-drop. Our overall conclusion from experiment and simulation is that the pressure-denatured state of λ*YA contains mainly residual helix and little β-sheet; following a fast P-drop, at least some λ*YA forms misplaced helical structure within microseconds. We hypothesize that nonnative helix at helix-turn interfaces traps the protein in compact nonnative conformations. These traps delay the folding of at least some of the population for 1.4 ms en route to the native state. Based on molecular dynamics, we predict specific mutations at the helix-turn interfaces that should speed up refolding from the pressure-denatured state, if this hypothesis is correct. PMID:23620522

  20. Folded membrane dialyzer with mechanically sealed edges

    DOEpatents

    Markley, Finley W.

    1976-01-01

    A semipermeable membrane is folded in accordion fashion to form a stack of pleats and the edges are sealed so as to isolate the opposite surfaces of the membrane. The stack is contained within a case that provides ports for flow of blood in contact with one surface of the membrane through channels formed by the pleats and also provides ports for flow of a dialysate through channels formed by the pleats in contact with the other surface of the membrane. The serpentine side edges of the membrane are sealed by a solidified plastic material, whereas effective mechanical means are provided to seal the end edges of the folded membrane. The mechanical means include a clamping strip which biases case sealing flanges into a sealed relationship with end portions of the membrane near the end edges, which portions extend from the stack and between the sealing flanges.

  1. Chevron folding patterns and heteroclinic orbits

    NASA Astrophysics Data System (ADS)

    Budd, Christopher J.; Chakhchoukh, Amine N.; Dodwell, Timothy J.; Kuske, Rachel

    2016-09-01

    We present a model of multilayer folding in which layers with bending stiffness EI are separated by a very stiff elastic medium of elasticity k2 and subject to a horizontal load P. By using a dynamical system analysis of the resulting fourth order equation, we show that as the end shortening per unit length E is increased, then if k2 is large there is a smooth transition from small amplitude sinusoidal solutions at moderate values of P to larger amplitude chevron folds, with straight limbs separated by regions of high curvature when P is large. The chevron solutions take the form of near heteroclinic connections in the phase-plane. By means of this analysis, values for P and the slope of the limbs are calculated in terms of E and k2.

  2. Ca-Dependent Folding of Human Calumenin.

    PubMed

    Mazzorana, Marco; Hussain, Rohanah; Sorensen, Thomas

    2016-01-01

    Human calumenin (hCALU) is a six EF-hand protein belonging to the CREC family. As other members of the family, it is localized in the secretory pathway and regulates the activity of SERCA2a and of the ryanodine receptor in the endoplasmic reticulum (ER). We have studied the effects of Ca2+ binding to the protein and found it to attain a more compact structure upon ion binding. Circular Dichroism (CD) measurements suggest a major rearrangement of the protein secondary structure, which reversibly switches from disordered at low Ca2+ concentrations to predominantly alpha-helical when Ca2+ is added. SAXS experiments confirm the transition from an unfolded to a compact structure, which matches the structural prediction of a trilobal fold. Overall our experiments suggest that calumenin is a Ca2+ sensor, which folds into a compact structure, capable of interacting with its molecular partners, when Ca2+ concentration within the ER reaches the millimolar range. PMID:26991433

  3. RNA Hairpin Folding in the Crowded Cell

    PubMed Central

    Gao, Mimi; Gnutt, David; Orban, Axel; Appel, Bettina; Righetti, Francesco; Winter, Roland; Narberhaus, Franz; Müller, Sabine

    2016-01-01

    Abstract Precise secondary and tertiary structure formation is critically important for the cellular functionality of ribonucleic acids (RNAs). RNA folding studies were mainly conducted in vitro, without the possibility of validating these experiments inside cells. Here, we directly resolve the folding stability of a hairpin‐structured RNA inside live mammalian cells. We find that the stability inside the cell is comparable to that in dilute physiological buffer. On the contrary, the addition of in vitro artificial crowding agents, with the exception of high‐molecular‐weight PEG, leads to a destabilization of the hairpin structure through surface interactions and reduction in water activity. We further show that RNA stability is highly variable within cell populations as well as within subcellular regions of the cytosol and nucleus. We conclude that inside cells the RNA is subject to (localized) stabilizing and destabilizing effects that lead to an on average only marginal modulation compared to diluted buffer. PMID:26833452

  4. Fast-Folding Proteins under Stress

    PubMed Central

    Dave, Kapil; Gruebele, Martin

    2015-01-01

    Proteins are subject to a variety of stresses in biological organisms, including pressure and temperature, which are the easiest stresses to simulate by molecular dynamics. We discuss the effect of pressure and thermal stress on very fast folding model proteins, whose in vitro folding can be fully simulated on computers and compared with experiments. We then discuss experiments that can be used to subject proteins to low and high temperature unfolding, as well as low and high pressure unfolding. Pressure and temperature are prototypical perturbations that illustrate how close many proteins are to instability, a property that cells can exploit to control protein function. We conclude by reviewing some recent in-cell experiments, and progress being made in simulating and measuring protein stability and function inside live cells. PMID:26231095

  5. Ca-Dependent Folding of Human Calumenin

    PubMed Central

    Mazzorana, Marco; Hussain, Rohanah; Sorensen, Thomas

    2016-01-01

    Human calumenin (hCALU) is a six EF-hand protein belonging to the CREC family. As other members of the family, it is localized in the secretory pathway and regulates the activity of SERCA2a and of the ryanodine receptor in the endoplasmic reticulum (ER). We have studied the effects of Ca2+ binding to the protein and found it to attain a more compact structure upon ion binding. Circular Dichroism (CD) measurements suggest a major rearrangement of the protein secondary structure, which reversibly switches from disordered at low Ca2+ concentrations to predominantly alpha-helical when Ca2+ is added. SAXS experiments confirm the transition from an unfolded to a compact structure, which matches the structural prediction of a trilobal fold. Overall our experiments suggest that calumenin is a Ca2+ sensor, which folds into a compact structure, capable of interacting with its molecular partners, when Ca2+ concentration within the ER reaches the millimolar range. PMID:26991433

  6. Predictive Computational Modeling of Chromatin Folding

    NASA Astrophysics Data System (ADS)

    di Pierro, Miichele; Zhang, Bin; Wolynes, Peter J.; Onuchic, Jose N.

    In vivo, the human genome folds into well-determined and conserved three-dimensional structures. The mechanism driving the folding process remains unknown. We report a theoretical model (MiChroM) for chromatin derived by using the maximum entropy principle. The proposed model allows Molecular Dynamics simulations of the genome using as input the classification of loci into chromatin types and the presence of binding sites of loop forming protein CTCF. The model was trained to reproduce the Hi-C map of chromosome 10 of human lymphoblastoid cells. With no additional tuning the model was able to predict accurately the Hi-C maps of chromosomes 1-22 for the same cell line. Simulations show unknotted chromosomes, phase separation of chromatin types and a preference of chromatin of type A to sit at the periphery of the chromosomes.

  7. RNA Hairpin Folding in the Crowded Cell.

    PubMed

    Gao, Mimi; Gnutt, David; Orban, Axel; Appel, Bettina; Righetti, Francesco; Winter, Roland; Narberhaus, Franz; Müller, Sabine; Ebbinghaus, Simon

    2016-02-24

    Precise secondary and tertiary structure formation is critically important for the cellular functionality of ribonucleic acids (RNAs). RNA folding studies were mainly conducted in vitro, without the possibility of validating these experiments inside cells. Here, we directly resolve the folding stability of a hairpin-structured RNA inside live mammalian cells. We find that the stability inside the cell is comparable to that in dilute physiological buffer. On the contrary, the addition of in vitro artificial crowding agents, with the exception of high-molecular-weight PEG, leads to a destabilization of the hairpin structure through surface interactions and reduction in water activity. We further show that RNA stability is highly variable within cell populations as well as within subcellular regions of the cytosol and nucleus. We conclude that inside cells the RNA is subject to (localized) stabilizing and destabilizing effects that lead to an on average only marginal modulation compared to diluted buffer.

  8. Chen’s Double Eyelid Fold Ratio

    PubMed Central

    Chen, Chen-Chia; Tai, Hao-Chih

    2016-01-01

    Background: Double eyelidplasty can construct palpebral folds and enhance beauty perception for Asians with single eyelids. A new palpebral parameter for the quantitative interpretation of surgical outcomes is proposed on the basis of a photometric study of the altered proportions of Asian eyes after double eyelid operation. Methods: A total of 100 Asian adults with single upper eyelids who were satisfied with the enlarged eyes by operation were included in the study. A retrospective measurement of palpebral parameters in the frontal profile both preoperatively and 6 months postoperatively was performed. The proportions of various parameters in the eyebrow–eye aesthetic unit were calculated and analyzed. Results: Double eyelidplasty can augment the vertical dimension of palpebral fissure by 27.9% increase on average. The vertical ratio of palpebral fissure to the eyebrow–eye unit is augmented by 34.4% increase. The vertical ratio of the subunit below double eyelid fold peak to the unit is augmented by 82.6% increase. Conclusions: Double eyelidplasty can substantially enlarge the vertical dimensions of the eyes of Asians with single eyelids. The eyes are perceived to be larger because of the visually assimilated illusion of the superimposed eyelid fold and the relative proportions of the eyebrow–eye unit. The authors propose using a vertical ratio of the subunit below double eyelid fold peak in the eyebrow–eye unit to measure the visually perceived proportion of the eye in the unit. This ratio can be applied clinically for a quantitative evaluation of the surgical outcome after double eyelidplasty. PMID:27200243

  9. Protein Folding Stages and Universal Exponents

    NASA Astrophysics Data System (ADS)

    Huang, Kerson

    We propose three stages in protein folding, based on physical arguements involving the interplay between the hydrophobic effect and hydrogen bonding, and computer simulations using the CSAW (conditioned self-avoiding walk) model. These stages are characterized by universal exponents ν = 3/5, 3/7, 2/5 in the power law R ~ Nν, where R is the radius of gyration and N is the number of residues. They correspond to the experimentally observed stages: unfolded, preglobule, molten globule.

  10. Protein Folding Stages and Universal Exponents

    NASA Astrophysics Data System (ADS)

    Huang, Kerson

    2011-11-01

    We propose three stages in protein folding, based on physical arguements involving the interplay between the hydrophobic effect and hydrogen bonding, and computer simulations using the CSAW (conditioned self-avoiding walk) model. These stages are characterized by universal exponents ν = 3/5, 3/7, 2/5 in the power law R ˜ Nν, where R is the radius of gyration and N is the number of residues. They correspond to the experimentally observed stages: unfolded, preglobule, molten globule.

  11. Understanding the Folding-Function Tradeoff in Proteins

    PubMed Central

    Gosavi, Shachi

    2013-01-01

    When an amino-acid sequence cannot be optimized for both folding and function, folding can get compromised in favor of function. To understand this tradeoff better, we devise a novel method for extracting the “function-less” folding-motif of a protein fold from a set of structurally similar but functionally diverse proteins. We then obtain the β-trefoil folding-motif, and study its folding using structure-based models and molecular dynamics simulations. CompariA protein sequence serves two purpson with the folding of wild-type β-trefoil proteins shows that function affects folding in two ways: In the slower folding interleukin-1β, binding sites make the fold more complex, increase contact order and slow folding. In the faster folding hisactophilin, residues which could have been part of the folding-motif are used for function. This reduces the density of native contacts in functional regions and increases folding rate. The folding-motif helps identify subtle structural deviations which perturb folding. These may then be used for functional annotation. Further, the folding-motif could potentially be used as a first step in the sequence design of function-less scaffold proteins. Desired function can then be engineered into these scaffolds. PMID:23593437

  12. Interaction Signatures Stabilizing the NAD(P)-Binding Rossmann Fold: A Structure Network Approach

    PubMed Central

    Bhattacharyya, Moitrayee; Upadhyay, Roopali; Vishveshwara, Saraswathi

    2012-01-01

    The fidelity of the folding pathways being encoded in the amino acid sequence is met with challenge in instances where proteins with no sequence homology, performing different functions and no apparent evolutionary linkage, adopt a similar fold. The problem stated otherwise is that a limited fold space is available to a repertoire of diverse sequences. The key question is what factors lead to the formation of a fold from diverse sequences. Here, with the NAD(P)-binding Rossmann fold domains as a case study and using the concepts of network theory, we have unveiled the consensus structural features that drive the formation of this fold. We have proposed a graph theoretic formalism to capture the structural details in terms of the conserved atomic interactions in global milieu, and hence extract the essential topological features from diverse sequences. A unified mathematical representation of the different structures together with a judicious concoction of several network parameters enabled us to probe into the structural features driving the adoption of the NAD(P)-binding Rossmann fold. The atomic interactions at key positions seem to be better conserved in proteins, as compared to the residues participating in these interactions. We propose a “spatial motif” and several “fold specific hot spots” that form the signature structural blueprints of the NAD(P)-binding Rossmann fold domain. Excellent agreement of our data with previous experimental and theoretical studies validates the robustness and validity of the approach. Additionally, comparison of our results with statistical coupling analysis (SCA) provides further support. The methodology proposed here is general and can be applied to similar problems of interest. PMID:23284738

  13. Resummation of semiclassical short folded string

    NASA Astrophysics Data System (ADS)

    Beccaria, Matteo; Macorini, Guido

    2012-02-01

    We reconsider semiclassical quantization of folded string spinning in AdS3 part of AdS5 × S5 using integrability-based (algebraic curve) method. We focus on the "short string" (small spin S) limit with the angular momentum J in S 5 scaled down according to {mathcal J} = ρ sqrt {S} in terms of the variables {mathcal J} = J/ sqrt {λ } , S = S/ sqrt {λ } . The semi-classical string energy in this particular scaling limit admits the double expansion E = {sum {_{{n = 0}}^{infty }sum {_{{p = 0}}^{infty }left( {sqrt {λ } } right)} }^{{1 - n}}}{a_{{n,p}}}left( ρ right){S^{{P + 1/2}}} . It behaves smoothly as J → 0 and partially resums recent results by Gromov and Valatka. We explicitly compute various one-loop coefficients a1, p ( ρ) by summing over the fluctuation frequencies for integrable perturbations around the classical solution. For the simple folded string, the result agrees with what could be derived exploiting a recent conjecture of Basso. However, the method can be extended to more general situations. As an example, we consider the m-folded string where Basso's conjecture fails. For this classical solution, we present the exact values of a 1,0( ρ) and a 1,1( ρ) for m = 2, 3, 4, 5 and explain how to work out the general case.

  14. Folding pathways of prion and doppel.

    PubMed Central

    Settanni, Giovanni; Hoang, Trinh Xuan; Micheletti, Cristian; Maritan, Amos

    2002-01-01

    The relevance of various residue positions for the stability and the folding characteristics of the prion protein in its normal cellular form are investigated by using molecular dynamics simulations of models exploiting the topology of the native state. These models allow for reproducing the experimentally validated two-state behavior of the normal prion isoform. Highly significant correlations are found between the most topologically relevant sites in our analysis and the single point mutations known to be associated with the arousal of the genetic forms of prion disease. Insight into the conformational change is provided by comparing the folding process of cellular prion and doppel that share a similar native state topology: the folding pathways of the former can be grouped in two main classes according to which tertiary structure contacts are formed first enroute to the native state. For the latter a single class of pathways leads to the native state again through a two-state process. Our results are consistent and supportive of the recent experimental findings that doppel lacks the scrapie isoform and that such remarkably different behavior involves residues in the region containing the two beta-strands and the intervening helix. PMID:12496120

  15. Simulating protein folding in different environmental conditions.

    PubMed

    Homouz, Dirar

    2014-01-01

    Molecular dynamics simulations have become an invaluable tool in investigating the dynamics of protein folding. However, most computational studies of protein folding assume dilute aqueous simulation conditions in order to reduce the complexity of the system under study and enhance the efficiency. Nowadays, it is evident that environmental conditions encountered in vivo (or even in vitro) play a major role in regulating the dynamics of protein folding especially when one considers the highly condensed environment in the cellular cytoplasm. In order to factor in these conditions, we can utilize the high efficiency of well-designed low resolution (coarse-grained) simulation models to reduce the complexity of these added protein-milieu interactions involving different time and length scales. The goal of this chapter is to describe some recently developed coarse-grained simulation techniques that are specifically designed to go beyond traditional aqueous solvent conditions. The chapter also gives the reader a flavor of the things that we can study using such "smart" low resolution models.

  16. Computing the conformational entropy for RNA folds

    NASA Astrophysics Data System (ADS)

    Liu, Liang; Chen, Shi-Jie

    2010-06-01

    We develop a polymer physics-based method to compute the conformational entropy for RNA tertiary folds, namely, conformations consisting of multiple helices connected through (cross-linked) loops. The theory is based on a virtual bond conformational model for the nucleotide chain. A key issue in the calculation of the entropy is how to treat the excluded volume interactions. The weak excluded volume interference between the different loops leads to the decomposition of the whole structure into a number of three-body building blocks, each consisting of a loop and two helices connected to the two ends of the loop. The simple construct of the three-body system allows an accurate computation for the conformational entropy for each building block. The assembly of the building blocks gives the entropy of the whole structure. This approach enables treatment of molten globule-like folds (partially unfolded tertiary structures) for RNAs. Extensive tests against experiments and exact computer enumerations indicate that the method can give accurate results for the entropy. The method developed here provides a solid first step toward a systematic development of a theory for the entropy and free energy landscape for complex tertiary folds for RNAs and proteins.

  17. RNA folding pathways in stop motion.

    PubMed

    Bottaro, Sandro; Gil-Ley, Alejandro; Bussi, Giovanni

    2016-07-01

    We introduce a method for predicting RNA folding pathways, with an application to the most important RNA tetraloops. The method is based on the idea that ensembles of three-dimensional fragments extracted from high-resolution crystal structures are heterogeneous enough to describe metastable as well as intermediate states. These ensembles are first validated by performing a quantitative comparison against available solution nuclear magnetic resonance (NMR) data of a set of RNA tetranucleotides. Notably, the agreement is better with respect to the one obtained by comparing NMR with extensive all-atom molecular dynamics simulations. We then propose a procedure based on diffusion maps and Markov models that makes it possible to obtain reaction pathways and their relative probabilities from fragment ensembles. This approach is applied to study the helix-to-loop folding pathway of all the tetraloops from the GNRA and UNCG families. The results give detailed insights into the folding mechanism that are compatible with available experimental data and clarify the role of intermediate states observed in previous simulation studies. The method is computationally inexpensive and can be used to study arbitrary conformational transitions. PMID:27091499

  18. Macrocyclization of Folded Diamines in Cavitands.

    PubMed

    Shi, Qixun; Masseroni, Daniele; Rebek, Julius

    2016-08-31

    Synthetic access to water-soluble cavitands and capsules has moved recognition events from organic solvents into aqueous media. Here we report the binding and reactivity of long-chain α,ω-diamines (C11 to C18) in cavitand hosts. The containers bind the diamines in folded conformations that bury the hydrocarbon chains and expose the amino groups to the aqueous medium. Their acylation with succinic anhydride results in improved yields of monofunctionalized products. The cavitand-bound amino acid products were cyclized to the corresponding macrocyclic dilactams in D2O using water-soluble carbodiimide. Direct reaction of the folded diamines in the cavitand with activated diesters of succinic acid and glutaric acids resulted in 54-96% yields of the 17- to 25-membered dilactams. These cavitand-chaperoned reactions provided 3- to 10-fold improvements over the yields obtained in bulk solution and offer an alternative to high dilution methods. The cavitand induces unlikely conformations in flexible guests and channels their reactivity along otherwise improbable paths. PMID:27529442

  19. Is Protein Folding Sub-Diffusive?

    PubMed Central

    Krivov, Sergei V.

    2010-01-01

    Protein folding dynamics is often described as diffusion on a free energy surface considered as a function of one or few reaction coordinates. However, a growing number of experiments and models show that, when projected onto a reaction coordinate, protein dynamics is sub-diffusive. This raises the question as to whether the conventionally used diffusive description of the dynamics is adequate. Here, we numerically construct the optimum reaction coordinate for a long equilibrium folding trajectory of a Go model of a -repressor protein. The trajectory projected onto this coordinate exhibits diffusive dynamics, while the dynamics of the same trajectory projected onto a sub-optimal reaction coordinate is sub-diffusive. We show that the higher the (cut-based) free energy profile for the putative reaction coordinate, the more diffusive the dynamics become when projected on this coordinate. The results suggest that whether the projected dynamics is diffusive or sub-diffusive depends on the chosen reaction coordinate. Protein folding can be described as diffusion on the free energy surface as function of the optimum reaction coordinate. And conversely, the conventional reaction coordinates, even though they might be based on physical intuition, are often sub-optimal and, hence, show sub-diffusive dynamics. PMID:20862361

  20. RNA folding pathways in stop motion

    PubMed Central

    Bottaro, Sandro; Gil-Ley, Alejandro; Bussi, Giovanni

    2016-01-01

    We introduce a method for predicting RNA folding pathways, with an application to the most important RNA tetraloops. The method is based on the idea that ensembles of three-dimensional fragments extracted from high-resolution crystal structures are heterogeneous enough to describe metastable as well as intermediate states. These ensembles are first validated by performing a quantitative comparison against available solution nuclear magnetic resonance (NMR) data of a set of RNA tetranucleotides. Notably, the agreement is better with respect to the one obtained by comparing NMR with extensive all-atom molecular dynamics simulations. We then propose a procedure based on diffusion maps and Markov models that makes it possible to obtain reaction pathways and their relative probabilities from fragment ensembles. This approach is applied to study the helix-to-loop folding pathway of all the tetraloops from the GNRA and UNCG families. The results give detailed insights into the folding mechanism that are compatible with available experimental data and clarify the role of intermediate states observed in previous simulation studies. The method is computationally inexpensive and can be used to study arbitrary conformational transitions. PMID:27091499

  1. Self-Folding Single Cell Grippers

    PubMed Central

    2015-01-01

    Given the heterogeneous nature of cultures, tumors, and tissues, the ability to capture, contain, and analyze single cells is important for genomics, proteomics, diagnostics, therapeutics, and surgery. Moreover, for surgical applications in small conduits in the body such as in the cardiovascular system, there is a need for tiny tools that approach the size of the single red blood cells that traverse the blood vessels and capillaries. We describe the fabrication of arrayed or untethered single cell grippers composed of biocompatible and bioresorbable silicon monoxide and silicon dioxide. The energy required to actuate these grippers is derived from the release of residual stress in 3–27 nm thick films, did not require any wires, tethers, or batteries, and resulted in folding angles over 100° with folding radii as small as 765 nm. We developed and applied a finite element model to predict these folding angles. Finally, we demonstrated the capture of live mouse fibroblast cells in an array of grippers and individual red blood cells in untethered grippers which could be released from the substrate to illustrate the potential utility for in vivo operations. PMID:24937214

  2. Cesium removal and kinetics equilibrium: Precipitation kinetics

    SciTech Connect

    Barnes, M.J.

    1999-12-17

    This task consisted of both non-radioactive and radioactive (tracer) tests examining the influence of potentially significant variables on cesium tetraphenylborate precipitation kinetics. The work investigated the time required to reach cesium decontamination and the conditions that affect the cesium precipitation kinetics.

  3. Center for Environmental Kinetic Synthesis (CEKA)

    SciTech Connect

    Lichtner, Peter .

    2006-06-01

    CEKA, as an Environmental Molecular Science Institute, is a joint research initiative of the National Science Foundation and U.S. Department of Energy, Biological and Environmental Research (BER). DOE collaborators are from DOE facilities at Los Alamos National Lab, Lawrence Berkeley National Lab and Pacific Northwest National Lab. The chief goals for CEKA are to 1) collect and synthesize molecular-level kinetic data into a coherent framework that can be used to predict time evolution of environmental processes over a range of temporal and spatial scales; 2) train a cohort of talented and diverse students to work on kinetic problems at multiple scales; 3) develop and promote the use of new experimental techniques in environmental kinetics; 4) develop and promote the use of new modeling tools to conceptualize reaction kinetics in environmental systems; and 5) communicate our understanding of issues related to environmental kinetics and issues of scale to the broader scientific community and to the public.

  4. Shear properties of vocal fold mucosal tissues and their effect on vocal fold oscillation

    NASA Astrophysics Data System (ADS)

    Chan, Roger Wai Kai

    Viscoelastic shear properties of vocal fold mucosal tissues and phonosurgical biomaterials were measured with a parallel-plate rotational rheometer. Elastic, viscous and damping properties were quantified as a function of frequency (0.01 Hz to 15 Hz) for human vocal fold mucosal tissues (N = 15), implantable biomaterials commonly used in the treatment of vocal fold paralysis (Teflon, gelatin, and collagen) (the non-mucosal group), and biomaterials currently or potentially useful in the treatment of vocal fold mucosal defects (adipose tissue or fat, hyaluronic acid, and fibronectin) (the mucosal group). It was found that intersubject differences as large as an order of magnitude were often observed for the shear properties of vocal fold mucosal tissues, part of which may be age- and gender-related. Shear properties of the non-mucosal group biomaterials were often much higher than those of the mucosal group biomaterials, which were relatively close to the shear properties of mucosal tissues. Viscoelastic and rheological modeling showed that shear properties of human vocal fold mucosa may be described by a quasi-linear viscoelastic theory and a statistical network theory, based upon which extrapolations to audio frequencies were possible. A theory of small-amplitude vocal fold oscillation was revisited to describe the effects of tissue shear properties on vocal fold oscillation and phonation threshold pressure, a measure of the 'ease' of phonation and an objective indication of vocal function. It was found that phonation threshold pressure is directly related to the viscous shear modulus or the 'effective damping modulus', a concept proposed to quantify the effective amount of damping in vocal fold oscillation. The mucosal group biomaterials were incorporated into the artificial vocal fold mucosa of a physical model in order to empirically assess their effects on phonation threshold pressure. Results showed that higher threshold pressures were consistently observed

  5. What makes a protein fold amenable to functional innovation? Fold polarity and stability trade-offs.

    PubMed

    Dellus-Gur, Eynat; Toth-Petroczy, Agnes; Elias, Mikael; Tawfik, Dan S

    2013-07-24

    Protein evolvability includes two elements--robustness (or neutrality, mutations having no effect) and innovability (mutations readily inducing new functions). How are these two conflicting demands bridged? Does the ability to bridge them relate to the observation that certain folds, such as TIM barrels, accommodate numerous functions, whereas other folds support only one? Here, we hypothesize that the key to innovability is polarity--an active site composed of flexible, loosely packed loops alongside a well-separated, highly ordered scaffold. We show that highly stabilized variants of TEM-1 β-lactamase exhibit selective rigidification of the enzyme's scaffold while the active-site loops maintained their conformational plasticity. Polarity therefore results in stabilizing, compensatory mutations not trading off, but instead promoting the acquisition of new activities. Indeed, computational analysis indicates that in folds that accommodate only one function throughout evolution, for example, dihydrofolate reductase, ≥ 60% of the active-site residues belong to the scaffold. In contrast, folds associated with multiple functions such as the TIM barrel show high scaffold-active-site polarity (~20% of the active site comprises scaffold residues) and >2-fold higher rates of sequence divergence at active-site positions. Our work suggests structural measures of fold polarity that appear to be correlated with innovability, thereby providing new insights regarding protein evolution, design, and engineering.

  6. Folding of β-barrel membrane proteins in lipid bilayers - Unassisted and assisted folding and insertion.

    PubMed

    Kleinschmidt, Jörg H

    2015-09-01

    In cells, β-barrel membrane proteins are transported in unfolded form to an outer membrane into which they fold and insert. Model systems have been established to investigate the mechanisms of insertion and folding of these versatile proteins into detergent micelles, lipid bilayers and even synthetic amphipathic polymers. In these experiments, insertion into lipid membranes is initiated from unfolded forms that do not display residual β-sheet secondary structure. These studies therefore have allowed the investigation of membrane protein folding and insertion in great detail. Folding of β-barrel membrane proteins into lipid bilayers has been monitored from unfolded forms by dilution of chaotropic denaturants that keep the protein unfolded as well as from unfolded forms present in complexes with molecular chaperones from cells. This review is aimed to provide an overview of the principles and mechanisms observed for the folding of β-barrel transmembrane proteins into lipid bilayers, the importance of lipid-protein interactions and the function of molecular chaperones and folding assistants. This article is part of a Special Issue entitled: Lipid-protein interactions.

  7. ProFold: Protein Fold Classification with Additional Structural Features and a Novel Ensemble Classifier

    PubMed Central

    2016-01-01

    Protein fold classification plays an important role in both protein functional analysis and drug design. The number of proteins in PDB is very large, but only a very small part is categorized and stored in the SCOPe database. Therefore, it is necessary to develop an efficient method for protein fold classification. In recent years, a variety of classification methods have been used in many protein fold classification studies. In this study, we propose a novel classification method called proFold. We import protein tertiary structure in the period of feature extraction and employ a novel ensemble strategy in the period of classifier training. Compared with existing similar ensemble classifiers using the same widely used dataset (DD-dataset), proFold achieves 76.2% overall accuracy. Another two commonly used datasets, EDD-dataset and TG-dataset, are also tested, of which the accuracies are 93.2% and 94.3%, higher than the existing methods. ProFold is available to the public as a web-server.

  8. ProFold: Protein Fold Classification with Additional Structural Features and a Novel Ensemble Classifier

    PubMed Central

    2016-01-01

    Protein fold classification plays an important role in both protein functional analysis and drug design. The number of proteins in PDB is very large, but only a very small part is categorized and stored in the SCOPe database. Therefore, it is necessary to develop an efficient method for protein fold classification. In recent years, a variety of classification methods have been used in many protein fold classification studies. In this study, we propose a novel classification method called proFold. We import protein tertiary structure in the period of feature extraction and employ a novel ensemble strategy in the period of classifier training. Compared with existing similar ensemble classifiers using the same widely used dataset (DD-dataset), proFold achieves 76.2% overall accuracy. Another two commonly used datasets, EDD-dataset and TG-dataset, are also tested, of which the accuracies are 93.2% and 94.3%, higher than the existing methods. ProFold is available to the public as a web-server. PMID:27660761

  9. Universality and diversity of folding mechanics for three-helix bundle proteins

    PubMed Central

    Yang, Jae Shick; Wallin, Stefan; Shakhnovich, Eugene I.

    2008-01-01

    In this study we evaluate, at full atomic detail, the folding processes of two small helical proteins, the B domain of protein A and the Villin headpiece. Folding kinetics are studied by performing a large number of ab initio Monte Carlo folding simulations using a single transferable all-atom potential. Using these trajectories, we examine the relaxation behavior, secondary structure formation, and transition-state ensembles (TSEs) of the two proteins and compare our results with experimental data and previous computational studies. To obtain a detailed structural information on the folding dynamics viewed as an ensemble process, we perform a clustering analysis procedure based on graph theory. Moreover, rigorous pfold analysis is used to obtain representative samples of the TSEs and a good quantitative agreement between experimental and simulated Φ values is obtained for protein A. Φ values for Villin also are obtained and left as predictions to be tested by future experiments. Our analysis shows that the two-helix hairpin is a common partially stable structural motif that gets formed before entering the TSE in the studied proteins. These results together with our earlier study of Engrailed Homeodomain and recent experimental studies provide a comprehensive, atomic-level picture of folding mechanics of three-helix bundle proteins. PMID:18195374

  10. Folding and imaging of DNA nanostructures in anhydrous and hydrated deep-eutectic solvents.

    PubMed

    Gállego, Isaac; Grover, Martha A; Hud, Nicholas V

    2015-06-01

    There is great interest in DNA nanotechnology, but its use has been limited to aqueous or substantially hydrated media. The first assembly of a DNA nanostructure in a water-free solvent, namely a low-volatility biocompatible deep-eutectic solvent composed of a 4:1 mixture of glycerol and choline chloride (glycholine), is now described. Glycholine allows for the folding of a two-dimensional DNA origami at 20 °C in six days, whereas in hydrated glycholine, folding is accelerated (≤3 h). Moreover, a three-dimensional DNA origami and a DNA tail system can be folded in hydrated glycholine under isothermal conditions. Glycholine apparently reduces the kinetic traps encountered during folding in aqueous solvent. Furthermore, folded structures can be transferred between aqueous solvent and glycholine. It is anticipated that glycholine and similar solvents will allow for the creation of functional DNA structures of greater complexity by providing a milieu with tunable properties that can be optimized for a range of applications and nanostructures.

  11. The Trigger Factor Chaperone Encapsulates and Stabilizes Partial Folds of Substrate Proteins

    PubMed Central

    Singhal, Kushagra; Vreede, Jocelyne; Mashaghi, Alireza; Tans, Sander J.; Bolhuis, Peter G.

    2015-01-01

    How chaperones interact with protein chains to assist in their folding is a central open question in biology. Obtaining atomistic insight is challenging in particular, given the transient nature of the chaperone-substrate complexes and the large system sizes. Recent single-molecule experiments have shown that the chaperone Trigger Factor (TF) not only binds unfolded protein chains, but can also guide protein chains to their native state by interacting with partially folded structures. Here, we used all-atom MD simulations to provide atomistic insights into how Trigger Factor achieves this chaperone function. Our results indicate a crucial role for the tips of the finger-like appendages of TF in the early interactions with both unfolded chains and partially folded structures. Unfolded chains are kinetically trapped when bound to TF, which suppresses the formation of transient, non-native end-to-end contacts. Mechanical flexibility allows TF to hold partially folded structures with two tips (in a pinching configuration), and to stabilize them by wrapping around its appendages. This encapsulation mechanism is distinct from that of chaperones such as GroEL, and allows folded structures of diverse size and composition to be protected from aggregation and misfolding interactions. The results suggest that an ATP cycle is not required to enable both encapsulation and liberation. PMID:26512985

  12. De novo prediction of protein folding pathways and structure using the principle of sequential stabilization

    PubMed Central

    Adhikari, Aashish N.; Freed, Karl F.; Sosnick, Tobin R.

    2012-01-01

    Motivated by the relationship between the folding mechanism and the native structure, we develop a unified approach for predicting folding pathways and tertiary structure using only the primary sequence as input. Simulations begin from a realistic unfolded state devoid of secondary structure and use a chain representation lacking explicit side chains, rendering the simulations many orders of magnitude faster than molecular dynamics simulations. The multiple round nature of the algorithm mimics the authentic folding process and tests the effectiveness of sequential stabilization (SS) as a search strategy wherein 2° structural elements add onto existing structures in a process of progressive learning and stabilization of structure found in prior rounds of folding. Because no a priori knowledge is used, we can identify kinetically significant non-native interactions and intermediates, sometimes generated by only two mutations, while the evolution of contact matrices is often consistent with experiments. Moreover, structure prediction improves substantially by incorporating information from prior rounds. The success of our simple, homology-free approach affirms the validity of our description of the primary determinants of folding pathways and structure, and the effectiveness of SS as a search strategy. PMID:23045636

  13. Protein folding trajectories can be described quantitatively by one-dimensional diffusion over measured energy landscapes

    NASA Astrophysics Data System (ADS)

    Neupane, Krishna; Manuel, Ajay P.; Woodside, Michael T.

    2016-07-01

    Protein folding features a diffusive search over a multidimensional energy landscape in conformational space for the minimum-energy structure. Experiments, however, are usually interpreted in terms of a one-dimensional (1D) projection of the full landscape onto a practical reaction coordinate. Although simulations have shown that folding kinetics can be described well by diffusion over a 1D projection, 1D approximations have not yet been fully validated experimentally. We used folding trajectories of single molecules held under tension in optical tweezers to compare the conditional probability of being on a transition path, calculated from the trajectory, with the prediction for ideal 1D diffusion over the measured 1D landscape, calculated from committor statistics. We found good agreement for the protein PrP (refs ,) and for one of the structural transitions in a leucine-zipper coiled-coil, but not for a second transition in the coiled-coil, owing to poor reaction-coordinate quality. These results show that 1D descriptions of folding can indeed be good, even for complex tertiary structures. More fundamentally, they also provide a fully experimental validation of the basic physical picture of folding as diffusion over a landscape.

  14. Efficient detection of secondary structure folded nucleic acids related to Alzheimer's disease based on junction probes.

    PubMed

    Li, Juan; Qi, Xiu-Juan; Du, Yan-Yan; Fu, Hua-E; Chen, Guo-Nan; Yang, Huang-Hao

    2012-01-01

    Single stranded DNA often forms stable secondary structures under physiological conditions. These DNA secondary structures play important physiological roles. However, the analysis of such secondary structure folded DNA is often complicated because of its high thermodynamic stability and slow hybridization kinetics. In this article, we demonstrate that Y-shaped junction probes could be used for rapid and highly efficient detection of secondary structure folded DNA. Our approach contained a molecular beacon (MB) probe and an assistant probe. In the absence of target, the MB probe failed to hybridize with the assistant probe. Whereas, the MB probe and the assistant probe could cooperatively unwind the secondary structure folded DNA target to form a ternary Y-shaped junction structure. In this condition, the MB probe was also opened, resulting in separating the fluorophores from the quenching moiety and emitting the fluorescence signal. This approach allowed for the highly sensitive detection of secondary structure folded DNA target, such as a tau specific DNA fragment related to Alzheimer's disease in this case. Additionally, this approach showed strong SNPs identifying capability. Furthermore, it was noteworthy that this newly proposed approach was capable of detecting secondary structure folded DNA target in cell lysate samples.

  15. Statics, metastable states, and barriers in protein folding: A replica variational approach

    NASA Astrophysics Data System (ADS)

    Takada, Shoji; Wolynes, Peter G.

    1997-04-01

    Protein folding is analyzed using a replica variational formalism to investigate some free energy landscape characteristics relevant for dynamics. A random contact interaction model that satisfies the minimum frustration principle is used to describe the coil-globule transition (characterized by TCG), glass transitions (by TA and TK), and folding transition (by TF). Trapping on the free energy landscape is characterized by two characteristic temperatures, one dynamic (TA) and the other static [TK (TA>TK)], which are similar to those found in mean field theories of the Potts glass. (i) Above TA, the free energy landscape is monotonous and the polymer is melted both dynamically and statically. (ii) Between TA and TK, the melted phase is still dominant thermodynamically, but frozen metastable states, exponentially large in number, appear. (iii) A few lowest minima become thermodynamically dominant below TK, where the polymer is totally frozen. In the temperature range between TA and TK, barriers between metastable states are shown to grow with decreasing temperature, suggesting super-Arrhenius behavior in a sufficiently large system. Due to evolutionary constraints on fast folding, the folding temperature TF is expected to be higher than TK, but may or may not be higher than TA. Diverse scenarios of the folding kinetics are discussed based on phase diagrams that take into account the dynamical transition, as well as the static ones.

  16. A Rat Excised Larynx Model of Vocal Fold Scar

    ERIC Educational Resources Information Center

    Welham, Nathan V.; Montequin, Douglas W.; Tateya, Ichiro; Tateya, Tomoko; Choi, Seong Hee; Bless, Diane M.

    2009-01-01

    Purpose: To develop and evaluate a rat excised larynx model for the measurement of acoustic, aerodynamic, and vocal fold vibratory changes resulting from vocal fold scar. Method: Twenty-four 4-month-old male Sprague-Dawley rats were assigned to 1 of 4 experimental groups: chronic vocal fold scar, chronic vocal fold scar treated with 100-ng basic…

  17. Effect of hydrophobic interactions on the folding mechanism of β-hairpins.

    PubMed

    Popp, Alexander; Wu, Ling; Keiderling, Timothy A; Hauser, Karin

    2014-12-11

    Hydrophobic interactions are essential in stabilizing protein structures. How they affect the folding pathway and kinetics, however, is less clear. We used time-resolved infrared spectroscopy to study the dynamics of hydrophobic interactions of β-hairpin variants of the sequence Trpzip2 (SWTWENGKWTWK-NH2) that is stabilized by two cross-strand Trp-Trp pairs. The hydrophobicity strength was varied by substituting the tryptophans pairwise by either tyrosines or valines. Relaxation dynamics were induced by a laser-excited temperature jump, which separately probed for the loss of the cross-strand β-hairpin interaction and the rise of the disordered structure. All substitutions tested result in reduced thermal stability, lower transition temperatures, and faster dynamics compared to Trpzip2. However, the changes in folding dynamics depend on the amino acid substituted for Trp. The aromatic substitution of Tyr for Trp results in the same kinetics for the unfolding of sheet and growth of disorder, with similar activation energies, independent of the substitution position. Substitution of Trp with a solely hydrophobic Val results in even faster kinetics than substitution with Tyr but is additionally site-dependent. If the hairpin has a Val pair close to its termini, the rate constants for loss of sheet and gain of disorder are the same, but if the pair is close to the turn, the sheet and disorder components show different relaxation kinetics. The Trp → Val substitutions reveal that hydrophobic interactions alone weakly stabilize the hairpin structure, but adding edge-to-face aromatic interaction strengthens it, and both modify the complex folding process.

  18. Experimental folding and boudinage under pure constrictional conditions

    NASA Astrophysics Data System (ADS)

    Kobberger, Gustav

    1995-07-01

    Constrictional folds are characterized by true fold-axis parallel extension if the rock-volume does not vary during deformation. Studies of such folds in experiments, using plasticine layers of different apparent viscosity and power-law exponent, clearly indicate that fold-axis parallel stretch may be accompanied by plastic elongation as well as boudinage of the competent layer. Characteristic aspects of the experimentally folded competent layers are: (1) coeval development of folds and boudins; (2) layer thickness not changing during deformation; (3) layer-parallel shortening in sections perpendicular to the fold (stretching) axis; (4) enlargement of the initial thickness of the competent layer results in increasing fold wavelength and decreasing number of boudins. The ratio of dominant wavelength to layer thickness of the constrictional folds can be described mathematically approximately by the equation developed for plane strain folding of power-law materials

  19. Machine Learning: How Much Does It Tell about Protein Folding Rates?

    PubMed Central

    Chen, Heng-Chang; Bogatyreva, Natalya S.; Filion, Guillaume J.; Ivankov, Dmitry N.

    2015-01-01

    The prediction of protein folding rates is a necessary step towards understanding the principles of protein folding. Due to the increasing amount of experimental data, numerous protein folding models and predictors of protein folding rates have been developed in the last decade. The problem has also attracted the attention of scientists from computational fields, which led to the publication of several machine learning-based models to predict the rate of protein folding. Some of them claim to predict the logarithm of protein folding rate with an accuracy greater than 90%. However, there are reasons to believe that such claims are exaggerated due to large fluctuations and overfitting of the estimates. When we confronted three selected published models with new data, we found a much lower predictive power than reported in the original publications. Overly optimistic predictive powers appear from violations of the basic principles of machine-learning. We highlight common misconceptions in the studies claiming excessive predictive power and propose to use learning curves as a safeguard against those mistakes. As an example, we show that the current amount of experimental data is insufficient to build a linear predictor of logarithms of folding rates based on protein amino acid composition. PMID:26606303

  20. Probing the Non-Native H Helix Translocation in Apomyoglobin Folding Intermediates

    PubMed Central

    2015-01-01

    Apomyoglobin folds via sequential helical intermediates that are formed by rapid collapse of the A, B, G, and H helix regions. An equilibrium molten globule with a similar structure is formed near pH 4. Previous studies suggested that the folding intermediates are kinetically trapped states in which folding is impeded by non-native packing of the G and H helices. Fluorescence spectra of mutant proteins in which cysteine residues were introduced at several positions in the G and H helices show differential quenching of W14 fluorescence, providing direct evidence of translocation of the H helix relative to helices A and G in both the kinetic and equilibrium intermediates. Förster resonance energy transfer measurements show that a 5-({2-[(acetyl)amino]ethyl}amino)naphthalene-1-sulfonic acid acceptor coupled to K140C (helix H) is closer to Trp14 (helix A) in the equilibrium molten globule than in the native state, by a distance that is consistent with sliding of the H helix in an N-terminal direction by approximately one helical turn. Formation of an S108C–L135C disulfide prevents H helix translocation in the equilibrium molten globule by locking the G and H helices into their native register. By enforcing nativelike packing of the A, G, and H helices, the disulfide resolves local energetic frustration and facilitates transient docking of the E helix region onto the hydrophobic core but has only a small effect on the refolding rate. The apomyoglobin folding landscape is highly rugged, with several energetic bottlenecks that frustrate folding; relief of any one of the major identified bottlenecks is insufficient to speed progression to the transition state. PMID:24857522