computational folding Dotaz Zobrazit nápovědu
Trojrozměrné struktury proteinů je možné předpovídat tak, že vezmeme denaturovaný protein a na počítači simulujeme proces jeho sbalení. Tento článek shrnuje úspěchy i úskalí tohoto postupu, zejména využití vysoce výkonných počítačů, projektů distribuovaných výpočtů, grafických karet a specializovaných počítačů.
The three-dimensional structure of a protein can be predicted by a simulation of its folding from fully denaturated state. This article review success stories as well as pitfalls of this approach, namely applications of high performance computers, distributed computing projects, graphical processing units and specialised hardware.
The structure of proteins as well as their folding/unfolding equilibrium are commonly attributed to H-bonding and hydrophobic interactions. We have used the molecular dynamic simulations in an explicit water environment based on the standard empirical potential as well as more accurately (and thus also more reliably) on the QM/MM potential. The simulations where the dispersion term was suppressed have led to a substantial change of the tryptophan-cage protein structure (unfolded structure). This structure cannot fold without the dispersion energy term, whereas, if it is covered fully, the system finds its native structure relatively quickly. This implies that after such physical factors as temperature and pH, the dispersion energy is an important factor in protein structure determination as well as in the protein folding/unfolding equilibrium. The loss of dispersion also affected the R-helical structure. On the other hand, weakening the electrostatic interactions (and thus H-bonding) affected the R-helical structure only to a minor extent.
To be biologically active, proteins must adopt specific folded three-dimensional tertiary structures. Yet the genetic information for the proteins specifies only the primary structure, the linear sequence of amino acids in the polypeptide backbone. Many purified proteins can spontaneously refold in vitro after being completely unfolded, so the three-dimensional structure must be determined by the primary structure. How this occurs has become known as the protein-folding problem. It was primarily of academic interest, but the advent of protein engineering and the ability to produce any protein, often in an insoluble, unfolded, inactive and useless form, has made it also of great practical importance. The problem can be divided into two main questions. The first is by what kinetic process or pathway does protein adopt its native and biologically active folded conformation and the other, what is the physical basis of the stability of folded conformations. The present review summarizes the present state of knowledge of the problem and attempts to put it into perspective for what could follows.
- MeSH
- biologické modely MeSH
- Creutzfeldtova-Jakobova nemoc diagnóza MeSH
- encefalopatie bovinní spongioformní diagnóza MeSH
- konformace proteinů MeSH
- lidé MeSH
- molekulární biologie MeSH
- molekulární konformace MeSH
- proteiny diagnostické užití ultrastruktura MeSH
- sekvence aminokyselin MeSH
- techniky in vitro MeSH
- termodynamika MeSH
- výpočetní biologie MeSH
- zobrazování trojrozměrné využití MeSH
- Check Tag
- lidé MeSH
We study the folding of the designed hairpin chignolin, using simulations with four different force fields. Interestingly, we find a misfolded, out-of-register, structure comprising 20-50% of the ordered structures with three force fields, but not with a fourth. A defining feature of the misfold is that Gly-7 adopts a β(PR) conformation rather than α(L). By reweighting, we show that differences between the force fields can mostly be attributed to differences in glycine properties. Benchmarking against NMR data suggests that the preference for β(PR) is not a force-field artifact. For chignolin, we show that including the misfold in the ensemble results in back-recalculated NMR observables in slightly better agreement with experiment than parameters calculated from a folded ensemble only. For comparison, we show by NMR and circular dichroism spectroscopy that the G7K mutant of chignolin, in which formation of this misfold is impossible, is well folded with stability similar to the wild-type and does not populate the misfolded state in simulation. Our results highlight the complexity of interpreting NMR data for small, weakly structured, peptides in solution, as well as the importance of accurate glycine parameters in force fields, for a correct description of turn structures.
The pyranose 2-oxidases from Trametes ochracea and Trametes pubescens share markedly similar amino acid sequences with identity of 93.4%. When expressed from the recombinant plasmids based on the same vector in the Escherichia coli host strain BL21(DE3) at higher growth temperatures, they differ strikingly in the formation of the inclusion bodies. Upon overexpression in the cultures performed at 28 degrees C, the specific activity of pyranose 2-oxidase from T. pubescens was eight times higher than that from T. ochracea: 93% of pyranose 2-oxidase from T. ochracea and only 15% of that from T. pubescens was present in the form of inclusion bodies. To ascertain the cause of this difference, both cloned genes were shuffled. Site-directed recombination of p2o cDNAs revealed that DNA constructs ending with 3' end of p2o cDNA from T. pubescens code for proteins that are folded into an active form to the greater extent, regardless of the gene expression level. "In silicio" analysis of physico-chemical properties of the protein sequences of pyranose 2-oxidases revealed that the sequence of amino acid residues 368-430, constituting the small, head domain of pyranose 2-oxidase from T. pubescens, affects positively the enzyme folding at higher cultivation temperatures. The domain differs in six amino acid residues from that of T. ochracea.
- MeSH
- Basidiomycota enzymologie MeSH
- buněčná inkluze metabolismus MeSH
- elektroforéza v polyakrylamidovém gelu MeSH
- enzymová indukce MeSH
- Escherichia coli MeSH
- financování organizované MeSH
- karbohydrátdehydrogenasy biosyntéza chemie metabolismus MeSH
- mutageneze cílená MeSH
- rekombinace genetická genetika MeSH
- sbalování proteinů MeSH
- sekvence aminokyselin MeSH
- subcelulární frakce metabolismus MeSH
- výpočetní biologie MeSH
- vztahy mezi strukturou a aktivitou MeSH
Purpose Phonations into a tube with the distal end either in the air or submerged in water are used for voice therapy. This study explores the effective mechanisms of these therapy methods. Method The study applied a physical model complemented by calculations from a computational model, and the results were compared to those that have been reported for humans. The effects of tube phonation on vocal tract resonances and oral pressure variation were studied. The relationships of transglottic pressure variation in time Ptrans ( t) versus glottal area variation in time GA( t) were constructed. Results The physical model revealed that, for the phonation on [u:] vowel through a glass resonance tube ending in the air, the 1st formant frequency ( F1 ) decreased by 67%, from 315 Hz to 105 Hz, thus slightly above the fundamental frequency ( F0 ) that was set to 90-94 Hz . For phonation through the tube into water, F1 decreased by 91%-92%, reaching 26-28 Hz, and the water bubbling frequency Fb ≅ 19-24 Hz was just below F1 . The relationships of Ptrans ( t) versus GA( t) clearly differentiate vowel phonation from both therapy methods, and show a physical background for voice therapy with tubes. It is shown that comparable results have been measured in humans during tube therapy. For the tube in air, F1 descends closer to F0 , whereas for the tube in water, the frequency Fb occurs close to the acoustic-mechanical resonance of the human vocal tract. Conclusion In both therapy methods, part of the airflow energy required for phonation is substituted by the acoustic energy utilizing the 1st acoustic resonance. Thus, less flow energy is needed for vocal fold vibration, which results in improved vocal efficiency. The effect can be stronger in water resistance therapy if the frequency Fb approaches the acoustic-mechanical resonance of the vocal tract, while simultaneously F0 is voluntarily changed close to F1.
- MeSH
- akustika řeči MeSH
- anatomické modely MeSH
- fonace fyziologie MeSH
- glottis MeSH
- hlasový trénink MeSH
- lidé MeSH
- plíce fyziologie MeSH
- počítačová simulace MeSH
- řečová terapie metody MeSH
- Check Tag
- lidé MeSH
- mužské pohlaví MeSH
- ženské pohlaví MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
The aeroacoustic mechanisms in human voice production are complex coupled processes that are still not fully understood. In this article, a hybrid numerical approach to analyzing sound generation in human voice production is presented. First, the fluid flow problem is solved using a parallel finite-volume computational fluid dynamics (CFD) solver on a fine computational mesh covering the larynx. The CFD simulations are run for four geometrical configurations: both with and without false vocal folds, and with fixed convergent or convergent-divergent motion of the medial vocal fold surface. Then the aeroacoustic sources and propagation of sound waves are calculated using Lighthill's analogy or acoustic perturbation equations on a coarse mesh covering the larynx, vocal tract, and radiation region near the mouth. Aeroacoustic sound sources are investigated in the time and frequency domains to determine their precise origin and correlation with the flow field. The problem of acoustic wave propagation from the larynx and vocal tract into the free field is solved using the finite-element method. Two different vocal-tract shapes are considered and modeled according to MRI vocal-tract data of the vowels /i/ and /u/. The spectra of the radiated sound evaluated from acoustic simulations show good agreement with formant frequencies known from human subjects.
- MeSH
- akustika * MeSH
- hlas * MeSH
- hlasové řasy fyziologie MeSH
- larynx fyziologie MeSH
- lidé MeSH
- vzduch * MeSH
- Check Tag
- lidé MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
Explicit solvent molecular dynamics simulations have been used to complement preceding experimental and computational studies of folding of guanine quadruplexes (G-DNA). We initiate early stages of unfolding of several G-DNAs by simulating them under no-salt conditions and then try to fold them back using standard excess salt simulations. There is a significant difference between G-DNAs with all-anti parallel stranded stems and those with stems containing mixtures of syn and anti guanosines. The most natural rearrangement for all-anti stems is a vertical mutual slippage of the strands. This leads to stems with reduced numbers of tetrads during unfolding and a reduction of strand slippage during refolding. The presence of syn nucleotides prevents mutual strand slippage; therefore, the antiparallel and hybrid quadruplexes initiate unfolding via separation of the individual strands. The simulations confirm the capability of G-DNA molecules to adopt numerous stable locally and globally misfolded structures. The key point for a proper individual folding attempt appears to be correct prior distribution of syn and anti nucleotides in all four G-strands. The results suggest that at the level of individual molecules, G-DNA folding is an extremely multi-pathway process that is slowed by numerous misfolding arrangements stabilized on highly variable timescales.
- MeSH
- DNA chemie MeSH
- G-kvadruplexy * MeSH
- jednovláknová DNA chemie MeSH
- lidé MeSH
- simulace molekulární dynamiky * MeSH
- telomery chemie MeSH
- Check Tag
- lidé MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
BACKGROUND: Guanine quadruplexes (GQs) play vital roles in many cellular processes and are of much interest as drug targets. In contrast to the availability of many structural studies, there is still limited knowledge on GQ folding. SCOPE OF REVIEW: We review recent molecular dynamics (MD) simulation studies of the folding of GQs, with an emphasis paid to the human telomeric DNA GQ. We explain the basic principles and limitations of all types of MD methods used to study unfolding and folding in a way accessible to non-specialists. We discuss the potential role of G-hairpin, G-triplex and alternative GQ intermediates in the folding process. We argue that, in general, folding of GQs is fundamentally different from funneled folding of small fast-folding proteins, and can be best described by a kinetic partitioning (KP) mechanism. KP is a competition between at least two (but often many) well-separated and structurally different conformational ensembles. MAJOR CONCLUSIONS: The KP mechanism is the only plausible way to explain experiments reporting long time-scales of GQ folding and the existence of long-lived sub-states. A significant part of the natural partitioning of the free energy landscape of GQs comes from the ability of the GQ-forming sequences to populate a large number of syn-anti patterns in their G-tracts. The extreme complexity of the KP of GQs typically prevents an appropriate description of the folding landscape using just a few order parameters or collective variables. GENERAL SIGNIFICANCE: We reconcile available computational and experimental studies of GQ folding and formulate basic principles characterizing GQ folding landscapes. This article is part of a Special Issue entitled "G-quadruplex" Guest Editor: Dr. Concetta Giancola and Dr. Daniela Montesarchio.
- MeSH
- denaturace nukleových kyselin MeSH
- DNA chemie MeSH
- G-kvadruplexy * MeSH
- guanin chemie MeSH
- kinetika MeSH
- lidé MeSH
- párování bází MeSH
- simulace molekulární dynamiky * MeSH
- telomery chemie MeSH
- vztahy mezi strukturou a aktivitou MeSH
- Check Tag
- lidé MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
- přehledy MeSH
Despite significant advances in the understanding of protein structure-function relationships, revealing protein folding pathways still poses a challenge due to a limited number of relevant experimental tools. Widely-used experimental techniques, such as calorimetry or spectroscopy, critically depend on a proper data analysis. Currently, there are only separate data analysis tools available for each type of experiment with a limited model selection. To address this problem, we have developed the CalFitter web server to be a unified platform for comprehensive data fitting and analysis of protein thermal denaturation data. The server allows simultaneous global data fitting using any combination of input data types and offers 12 protein unfolding pathway models for selection, including irreversible transitions often missing from other tools. The data fitting produces optimal parameter values, their confidence intervals, and statistical information to define unfolding pathways. The server provides an interactive and easy-to-use interface that allows users to directly analyse input datasets and simulate modelled output based on the model parameters. CalFitter web server is available free at https://loschmidt.chemi.muni.cz/calfitter/.
