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1.
Nature ; 575(7781): 180-184, 2019 11.
Artigo em Inglês | MEDLINE | ID: mdl-31695210

RESUMO

Methane is a powerful greenhouse gas and is targeted for emissions mitigation by the US state of California and other jurisdictions worldwide1,2. Unique opportunities for mitigation are presented by point-source emitters-surface features or infrastructure components that are typically less than 10 metres in diameter and emit plumes of highly concentrated methane3. However, data on point-source emissions are sparse and typically lack sufficient spatial and temporal resolution to guide their mitigation and to accurately assess their magnitude4. Here we survey more than 272,000 infrastructure elements in California using an airborne imaging spectrometer that can rapidly map methane plumes5-7. We conduct five campaigns over several months from 2016 to 2018, spanning the oil and gas, manure-management and waste-management sectors, resulting in the detection, geolocation and quantification of emissions from 564 strong methane point sources. Our remote sensing approach enables the rapid and repeated assessment of large areas at high spatial resolution for a poorly characterized population of methane emitters that often appear intermittently and stochastically. We estimate net methane point-source emissions in California to be 0.618 teragrams per year (95 per cent confidence interval 0.523-0.725), equivalent to 34-46 per cent of the state's methane inventory8 for 2016. Methane 'super-emitter' activity occurs in every sector surveyed, with 10 per cent of point sources contributing roughly 60 per cent of point-source emissions-consistent with a study of the US Four Corners region that had a different sectoral mix9. The largest methane emitters in California are a subset of landfills, which exhibit persistent anomalous activity. Methane point-source emissions in California are dominated by landfills (41 per cent), followed by dairies (26 per cent) and the oil and gas sector (26 per cent). Our data have enabled the identification of the 0.2 per cent of California's infrastructure that is responsible for these emissions. Sharing these data with collaborating infrastructure operators has led to the mitigation of anomalous methane-emission activity10.


Assuntos
Monitoramento Ambiental , Metano/análise , Gerenciamento de Resíduos , California , Efeito Estufa , Esterco , Metano/química , Metano/metabolismo , Gás Natural , Indústria de Petróleo e Gás/métodos , Petróleo , Águas Residuárias
2.
PLoS Genet ; 14(7): e1007417, 2018 07.
Artigo em Inglês | MEDLINE | ID: mdl-30024879

RESUMO

Cell death plays a major role during C. elegans oogenesis, where over half of the oogenic germ cells die in a process termed physiological apoptosis. How germ cells are selected for physiological apoptosis, or instead become oocytes, is not understood. Most oocytes produce viable embryos when apoptosis is blocked, suggesting that physiological apoptosis does not function to cull defective germ cells. Instead, cells targeted for apoptosis may function as nurse cells; the germline is syncytial, and all germ cells appear to contribute cytoplasm to developing oocytes. C. elegans has been a leading model for the genetics and molecular biology of apoptosis and phagocytosis, but comparatively few studies have examined the cell biology of apoptotic cells. We used live imaging to identify and examine pre-apoptotic germ cells in the adult gonad. After initiating apoptosis, germ cells selectively export their mitochondria into the shared pool of syncytial cytoplasm; this transport appears to use the microtubule motor kinesin. The apoptotic cells then shrink as they expel most of their remaining cytoplasm, and close off from the syncytium. Shortly thereafter the apoptotic cells restructure their microtubule and actin cytoskeletons, possibly to maintain cell integrity; the microtubules form a novel, cortical array of stabilized microtubules, and actin and cofilin organize into giant cofilin-actin rods. We discovered that some apoptotic germ cells are binucleate; the binucleate germ cells can develop into binucleate oocytes in apoptosis-defective strains, and appear capable of producing triploid offspring. Our results suggest that the nuclear layer of the germline syncytium becomes folded during mitosis and growth, and that binucleate cells arise as the layer unfolds or everts; all of the binucleate cells are subsequently removed by apoptosis. These results show that physiological apoptosis targets at least two distinct populations of germ cells, and that the apoptosis machinery efficiently recognizes cells with two nuclei.


Assuntos
Apoptose/fisiologia , Caenorhabditis elegans/fisiologia , Núcleo Celular/patologia , Células Germinativas/fisiologia , Microtúbulos/fisiologia , Animais , Animais Geneticamente Modificados , Proteínas de Caenorhabditis elegans/genética , Caspases/genética , Núcleo Celular/fisiologia , Citoplasma/fisiologia , Feminino , Masculino , Oócitos/fisiologia , Oogênese/fisiologia , Ovário/citologia , Ovário/fisiologia
3.
Otolaryngol Head Neck Surg ; 159(2): 300-302, 2018 08.
Artigo em Inglês | MEDLINE | ID: mdl-29558244

RESUMO

Intraoperative identification of the spinal accessory nerve (SAN) is key in reducing nerve injury. This study aims to explore the surgical anatomy of the SAN and 2 landmarks for its identification-the sternocleidomastoid branch of the occipital artery (SBOA) and superior sternocleidomastoid tendon (SST)-to propose a novel method of identifying the SAN during surgical neck dissections. Twelve cadavers underwent bilateral level II-V neck dissection identifying the SAN, SBOA, and SST. Variation was documented and distance between landmarks and the SAN measured. The most common arrangement had the SST most superficially followed by the SBOA and then the SAN. The SAN was 3.63 ± 4.02 mm from the artery and 2.31 ± 1.72 mm from the tendon. A triangle-bordered by the tendon laterally, artery medially, and digastric muscle superiorly-contained the SAN in 95.8% of cases. This relationship translated into a reliable technique to identify the SAN intraoperatively, which has been used successfully in practice.


Assuntos
Nervo Acessório/anatomia & histologia , Pontos de Referência Anatômicos , Esvaziamento Cervical/métodos , Músculos do Pescoço/inervação , Cadáver , Humanos
4.
Opt Express ; 25(8): 9186-9195, 2017 Apr 17.
Artigo em Inglês | MEDLINE | ID: mdl-28437992

RESUMO

The intrinsic spectral dimensionality indicates the observable degrees of freedom in Earth's solar-reflected light field, quantifying the diversity of spectral content accessible by visible and infrared remote sensing. The solar-reflected regime spans the 0.38 - 2.5 µm interval, and is captured by a wide range of current and planned instruments on both airborne and orbital platforms. To date there has been no systematic study of its spectral dimensionality as a function of space, time, and land cover. Here we report a multi-site, multi-year statistical survey by NASA's "Classic" Airborne Visible Near InfraRed Spectrometer (AVIRIS-C). AVIRIS-C measured large regions of California, USA, spanning wide latitudinal and elevation gradients containing all canonical MODIS land cover types. The spectral uniformity of the AVIRIS-C design enabled consistent in-scene assessment of measurement noise across acquisitions. The estimated dimensionality as a function of cover type ranged from the low 20s to the high 40s, and was approximately 50 for the combined dataset. This result indicates the high diversity of physical processes distinguishable by imaging spectrometers like AVIRIS-C for one region of the Earth.

5.
Protein Sci ; 25(1): 19-29, 2016 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-26266489

RESUMO

Molecular dynamics (MD) simulation is a well-established tool for the computational study of protein structure and dynamics, but its application to the important problem of protein structure prediction remains challenging, in part because extremely long timescales can be required to reach the native structure. Here, we examine the extent to which the use of low-resolution information in the form of residue-residue contacts, which can often be inferred from bioinformatics or experimental studies, can accelerate the determination of protein structure in simulation. We incorporated sets of 62, 31, or 15 contact-based restraints in MD simulations of ubiquitin, a benchmark system known to fold to the native state on the millisecond timescale in unrestrained simulations. One-third of the restrained simulations folded to the native state within a few tens of microseconds-a speedup of over an order of magnitude compared with unrestrained simulations and a demonstration of the potential for limited amounts of structural information to accelerate structure determination. Almost all of the remaining ubiquitin simulations reached near-native conformations within a few tens of microseconds, but remained trapped there, apparently due to the restraints. We discuss potential methodological improvements that would facilitate escape from these near-native traps and allow more simulations to quickly reach the native state. Finally, using a target from the Critical Assessment of protein Structure Prediction (CASP) experiment, we show that distance restraints can improve simulation accuracy: In our simulations, restraints stabilized the native state of the protein, enabling a reasonable structural model to be inferred.


Assuntos
Simulação de Dinâmica Molecular , Ubiquitina/química , Conformação Proteica , Dobramento de Proteína
6.
Perspect Med Educ ; 4(5): 268-71, 2015 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-26358978

RESUMO

With the increasing popularity and scale of peer teaching, it is imperative to develop methods that ensure the quality of teaching provided by undergraduate students. We used an established faculty development and quality assurance process in a novel context: peer observation of teaching for undergraduate peer tutors. We have developed a form to record observations and aid the facilitation of feedback. In addition, experienced peer tutors have been trained to observe peer-taught sessions and provide tutors with verbal and written feedback. We have found peer observation of teaching to be a feasible and acceptable process for improving quality of teaching provided by undergraduate medical students. However, feedback regarding the quality of peer observer's feedback may help to develop students' abilities further.

7.
Eukaryot Cell ; 14(9): 858-67, 2015 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-26092920

RESUMO

The gametogenesis program of the budding yeast Saccharomyces cerevisiae, also known as sporulation, employs unusual internal meiotic divisions, after which all four meiotic products differentiate within the parental cell. We showed previously that sporulation is typically accompanied by the destruction of discarded immature meiotic products through their exposure to proteases released from the mother cell vacuole, which undergoes an apparent programmed rupture. Here we demonstrate that vacuolar rupture contributes to de facto programmed cell death (PCD) of the meiotic mother cell itself. Meiotic mother cell PCD is accompanied by an accumulation of depolarized mitochondria, organelle swelling, altered plasma membrane characteristics, and cytoplasmic clearance. To ensure that the gametes survive the destructive consequences of developing within a cell that is executing PCD, we hypothesized that PCD is restrained from occurring until spores have attained a threshold degree of differentiation. Consistent with this hypothesis, gene deletions that perturb all but the most terminal postmeiotic spore developmental stages are associated with altered PCD. In these mutants, meiotic mother cells exhibit a delay in vacuolar rupture and then appear to undergo an alternative form of PCD associated with catastrophic consequences for the underdeveloped spores. Our findings reveal yeast sporulation as a context of bona fide PCD that is developmentally coordinated with gamete differentiation.


Assuntos
Apoptose , Meiose , Saccharomyces cerevisiae/fisiologia , Esporos Fúngicos/fisiologia , Deleção de Genes , Potencial da Membrana Mitocondrial , Saccharomyces cerevisiae/citologia , Saccharomyces cerevisiae/genética , Esporos Fúngicos/citologia , Esporos Fúngicos/genética
8.
Appl Opt ; 53(7): 1363-80, 2014 Mar 01.
Artigo em Inglês | MEDLINE | ID: mdl-24663366

RESUMO

The design, characteristics, and first test flight results are described of the Portable Remote Imaging Spectrometer, an airborne sensor specifically designed to address the challenges of coastal ocean remote sensing. The sensor incorporates several technologies that are demonstrated for the first time, to the best of our knowledge, in a working system in order to achieve a high performance level in terms of uniformity, signal-to-noise ratio, low polarization sensitivity, low stray light, and high spatial resolution. The instrument covers the 350-1050 nm spectral range with a 2.83 nm sampling per pixel, and a 0.88 mrad instantaneous field of view, with 608 cross-track pixels in a pushbroom configuration. Two additional infrared channels (1240 and 1610 nm) are measured by a spot radiometer housed in the same head. The spectrometer design is based on an optically fast (F/1.8) Dyson design form coupled to a wide angle two-mirror telescope in a configuration that minimizes polarization sensitivity without the use of a depolarizer. A grating with minimum polarization sensitivity and broadband efficiency was fabricated as well as a slit assembly with black (etched) silicon surface to minimize backscatter. First flight results over calibration sites as well as Monterey Bay in California have demonstrated good agreement between in situ and remotely sensed data, confirming the potential value of the sensor to the coastal ocean science community.


Assuntos
Aeronaves/instrumentação , Monitoramento Ambiental/instrumentação , Tecnologia de Sensoriamento Remoto/instrumentação , Água do Mar/análise , Água do Mar/química , Análise Espectral/instrumentação , Desenho de Equipamento , Análise de Falha de Equipamento , Miniaturização , Oceanos e Mares , Projetos Piloto
9.
J Chem Phys ; 139(16): 164106, 2013 Oct 28.
Artigo em Inglês | MEDLINE | ID: mdl-24182003

RESUMO

In molecular dynamics simulations, control over temperature and pressure is typically achieved by augmenting the original system with additional dynamical variables to create a thermostat and a barostat, respectively. These variables generally evolve on timescales much longer than those of particle motion, but typical integrator implementations update the additional variables along with the particle positions and momenta at each time step. We present a framework that replaces the traditional integration procedure with separate barostat, thermostat, and Newtonian particle motion updates, allowing thermostat and barostat updates to be applied infrequently. Such infrequent updates provide a particularly substantial performance advantage for simulations parallelized across many computer processors, because thermostat and barostat updates typically require communication among all processors. Infrequent updates can also improve accuracy by alleviating certain sources of error associated with limited-precision arithmetic. In addition, separating the barostat, thermostat, and particle motion update steps reduces certain truncation errors, bringing the time-average pressure closer to its target value. Finally, this framework, which we have implemented on both general-purpose and special-purpose hardware, reduces software complexity and improves software modularity.


Assuntos
Simulação de Dinâmica Molecular , Pressão , Temperatura , Artefatos
10.
J Phys Chem B ; 117(42): 12898-907, 2013 Oct 24.
Artigo em Inglês | MEDLINE | ID: mdl-23841719

RESUMO

Understanding the nature of the glass transition--the dramatic slowing of dynamics and eventual emergence of a disordered solid from a cooling liquid--is a fundamental challenge in physical science. A central characteristic of glass-forming liquids is a non-exponential main relaxation process. The extent of deviation from exponential relaxation typically becomes more pronounced on cooling. Theories that predict a growth of spatially heterogeneous dynamics as temperature is lowered can explain these observations. In apparent contradiction to these theories, however, some experiments suggest that certain substances--notably including the intensely studied molecular glass-former ortho-terphenyl (OTP)--have a main relaxation process whose shape is essentially temperature independent, even though other observables predicted to be correlated with the degree of dynamical heterogeneity are temperature dependent. Here we report the first simulations based on an atomistic model of OTP that reach equilibrium at temperatures well into the supercooled regime. We first show that the results of these simulations are in reasonable quantitative agreement with experimental data for several basic properties over a wide range of temperatures. We then focus on rotational relaxation, finding nearly exponential behavior at high temperatures with clearly increasing deviations as temperature is lowered. The much weaker temperature dependence observed in light-scattering experiments also emerges from the same simulation data when we calculate correlation functions similar to those probed experimentally; this highlights the diversity of temperature dependencies that can be obtained with different probes. Further analysis suggests that the temperature insensitivity observed in the light-scattering experiments stems from the dependence of these measurements on internal as well as rotational molecular motion. Within the temperature range of our OTP simulations, our results strongly suggest that this archetypal glass-former behaves as anticipated by theories of the glass transition that predict increasing non-exponentiality with cooling, and our simulations thus strengthen the evidence supporting such theories.


Assuntos
Compostos de Terfenil/química , Simulação de Dinâmica Molecular , Rotação , Temperatura , Termodinâmica
11.
J Gen Physiol ; 141(5): 619-32, 2013 May.
Artigo em Inglês | MEDLINE | ID: mdl-23589581

RESUMO

The difficulty in characterizing ion conduction through membrane channels at the level of individual permeation events has made it challenging to elucidate the mechanistic principles underpinning this fundamental physiological process. Using long, all-atom simulations enabled by special-purpose hardware, we studied K(+) permeation across the KV1.2/2.1 voltage-gated potassium channel. At experimentally accessible voltages, which include the physiological range, the simulated permeation rate was substantially lower than the experimentally observed rate. The current-voltage relationship was also nonlinear but became linear at much higher voltages. We observed permeation consistent with a "knock-on" mechanism at all voltages. At high voltages, the permeation rate was in accordance with our previously reported KV1.2 pore-only simulations, after the simulated voltages from the previous study were recalculated using the correct method, new insight into which is provided here. Including the voltage-sensing domains in the simulated channel brought the linear current-voltage regime closer to the experimentally accessible voltages. The simulated permeation rate, however, still underestimated the experimental rate, because formation of the knock-on intermediate occurred too infrequently. Reducing the interaction strength between the ion and the selectivity filter did not increase conductance. In complementary simulations of gramicidin A, similar changes in interaction strength did increase the observed permeation rate. Permeation nevertheless remained substantially below the experimental value, largely because of infrequent ion recruitment into the pore lumen. Despite the need to apply large voltages to simulate the permeation process, the apparent voltage insensitivity of the permeation mechanism suggests that the direct simulation of permeation at the single-ion level can provide fundamental physiological insight into ion channel function. Notably, our simulations suggest that the knock-on permeation mechanisms in KV1.2 and KcsA may be different.


Assuntos
Ativação do Canal Iônico/fisiologia , Canal de Potássio Kv1.2/metabolismo , Canais de Potássio Shab/metabolismo , Condutividade Elétrica , Potenciais da Membrana/fisiologia , Modelos Biológicos , Permeabilidade , Potássio/metabolismo
12.
Cell ; 152(3): 557-69, 2013 Jan 31.
Artigo em Inglês | MEDLINE | ID: mdl-23374350

RESUMO

Dimerization-driven activation of the intracellular kinase domains of the epidermal growth factor receptor (EGFR) upon extracellular ligand binding is crucial to cellular pathways regulating proliferation, migration, and differentiation. Inactive EGFR can exist as both monomers and dimers, suggesting that the mechanism regulating EGFR activity may be subtle. The membrane itself may play a role but creates substantial difficulties for structural studies. Our molecular dynamics simulations of membrane-embedded EGFR suggest that, in ligand-bound dimers, the extracellular domains assume conformations favoring dimerization of the transmembrane helices near their N termini, dimerization of the juxtamembrane segments, and formation of asymmetric (active) kinase dimers. In ligand-free dimers, by holding apart the N termini of the transmembrane helices, the extracellular domains instead favor C-terminal dimerization of the transmembrane helices, juxtamembrane segment dissociation and membrane burial, and formation of symmetric (inactive) kinase dimers. Electrostatic interactions of EGFR's intracellular module with the membrane are critical in maintaining this coupling.


Assuntos
Membrana Celular/metabolismo , Receptores ErbB/química , Membrana Celular/química , Dimerização , Receptores ErbB/metabolismo , Humanos , Lipídeos de Membrana/metabolismo , Simulação de Dinâmica Molecular , Ressonância Magnética Nuclear Biomolecular , Conformação Proteica , Estrutura Terciária de Proteína , Eletricidade Estática
13.
Autophagy ; 9(2): 263-5, 2013 Feb 01.
Artigo em Inglês | MEDLINE | ID: mdl-23187615

RESUMO

Studies of the budding yeast Saccharomyces cerevisiae have provided many of the most important insights into the mechanisms of autophagy, which are common to all eukaryotes. However, investigation of yeast self-destruction pathways, including autophagy and programmed cell death, has been almost exclusively restricted to cells undergoing vegetative growth, leaving very little exploration of their functions during developmental transitions in the yeast life cycle. We have recently discovered that whole nuclei are subject to programmed destruction during yeast gametogenesis. Programmed nuclear destruction (PND) possesses characteristics of apoptosis in the form of DNA cleavage by endonuclease G, and involves bulk protein turnover through an unusual autophagic pathway involving lysis of the vacuole rather than delivery of components to it through macroautophagy. We thus illuminate an example of developmentally programmed cellular "self-eating" in yeast, which is associated with the rupture of a lytic organelle, reminiscent of programmed cell death mechanisms in plants and animals.


Assuntos
Autofagia , Núcleo Celular/metabolismo , Saccharomyces cerevisiae/citologia , Saccharomyces cerevisiae/metabolismo , Vacúolos/metabolismo , Animais , Núcleo Celular/ultraestrutura , Saccharomyces cerevisiae/ultraestrutura , Proteínas de Saccharomyces cerevisiae/metabolismo , Esporos Fúngicos/citologia , Esporos Fúngicos/ultraestrutura
14.
Small ; 9(7): 1058-65, 2013 Apr 08.
Artigo em Inglês | MEDLINE | ID: mdl-23233398

RESUMO

The optical absorption efficiencies of vertically aligned multi-walled (MW)-carbon nanotube (CNT) ensembles are characterized in the 350-7000 nm wavelength range where CNT site densities > 1 × 10(11) /cm(2) are achieved directly on metallic substrates. The site density directly impacts the optical absorption characteristics, and while high-density arrays of CNTs on electrically insulating and non-metallic substrates have been commonly reported, achieving high site-densities on metals has been challenging and remains an area of active research. These absorber ensembles are ultra-thin (<10 µm) and yet they still exhibit a reflectance as low as ∼0.02%, which is 100 times lower than the reference; these characteristics make them potentially attractive for high-sensitivity and high-speed thermal detectors. In addition, the use of a plasma-enhanced chemical vapor deposition process for the synthesis of the absorbers increases the portfolio of materials that can be integrated with such absorbers due to the potential for reduced synthesis temperatures. The remarkable ruggedness of the absorbers is also demonstrated as they are exposed to high temperatures in an oxidizing ambient environment, making them well-suited for extreme thermal environments encountered in the field, potentially for solar cell applications. Finally, a phenomenological model enables the determinatiom of the extinction coefficients in these nanostructures and the results compare well with experiment.


Assuntos
Luz , Nanotecnologia/métodos , Nanotubos de Carbono/química , Microscopia de Força Atômica , Modelos Teóricos , Nanotubos de Carbono/ultraestrutura , Raios Ultravioleta
15.
Dev Cell ; 23(1): 35-44, 2012 Jul 17.
Artigo em Inglês | MEDLINE | ID: mdl-22727375

RESUMO

Autophagy controls cellular catabolism in diverse eukaryotes and modulates programmed cell death in plants and animals. While studies of the unicellular yeast Saccharomyces cerevisiae have provided fundamental insights into the mechanisms of autophagy, the roles of cell death pathways in yeast are less well understood. Here, we describe widespread developmentally programmed nuclear destruction (PND) events that occur during yeast gametogenesis. PND is executed through apoptotic-like DNA fragmentation in coordination with an unusual form of autophagy that is most similar to mammalian lysosomal membrane permeabilization and mega-autophagy, a form of plant autophagic cell death. Undomesticated strains execute gametogenic PND broadly in maturing colonies to the apparent benefit of sibling cells, confirming its prominence during the yeast life cycle. Our results reveal that diverse cell-death-related processes converge during gametogenesis in a microbe distantly related to plants or animals, highlighting gametogenesis as a process during which programmed cell death mechanisms may have evolved.


Assuntos
Núcleo Celular/fisiologia , Fragmentação do DNA , Gametogênese/fisiologia , Saccharomyces cerevisiae/crescimento & desenvolvimento , Saccharomyces cerevisiae/fisiologia , Esporos Fúngicos/fisiologia , Animais , Apoptose/fisiologia , Autofagia/fisiologia , Núcleo Celular/ultraestrutura , Gametogênese Vegetal/fisiologia , Lisossomos/fisiologia , Mamíferos , Saccharomyces cerevisiae/citologia , Esporos Fúngicos/ultraestrutura
16.
Cell ; 149(4): 860-70, 2012 May 11.
Artigo em Inglês | MEDLINE | ID: mdl-22579287

RESUMO

The mutation and overexpression of the epidermal growth factor receptor (EGFR) are associated with the development of a variety of cancers, making this prototypical dimerization-activated receptor tyrosine kinase a prominent target of cancer drugs. Using long-timescale molecular dynamics simulations, we find that the N lobe dimerization interface of the wild-type EGFR kinase domain is intrinsically disordered and that it becomes ordered only upon dimerization. Our simulations suggest, moreover, that some cancer-linked mutations distal to the dimerization interface, particularly the widespread L834R mutation (also referred to as L858R), facilitate EGFR dimerization by suppressing this local disorder. Corroborating these findings, our biophysical experiments and kinase enzymatic assays indicate that the L834R mutation causes abnormally high activity primarily by promoting EGFR dimerization rather than by allowing activation without dimerization. We also find that phosphorylation of EGFR kinase domain at Tyr845 may suppress the intrinsic disorder, suggesting a molecular mechanism for autonomous EGFR signaling.


Assuntos
Receptores ErbB/química , Receptores ErbB/genética , Neoplasias/metabolismo , Mutação Puntual , Transdução de Sinais , Sequência de Aminoácidos , Cristalografia por Raios X , Receptores ErbB/antagonistas & inibidores , Receptores ErbB/metabolismo , Gefitinibe , Humanos , Lapatinib , Simulação de Dinâmica Molecular , Dados de Sequência Molecular , Dobramento de Proteína , Inibidores de Proteínas Quinases/farmacologia , Multimerização Proteica , Estrutura Terciária de Proteína , Quinazolinas/farmacologia , Alinhamento de Sequência
17.
Proteins ; 80(8): 2071-9, 2012 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-22513870

RESUMO

Accurate computational prediction of protein structure represents a longstanding challenge in molecular biology and structure-based drug design. Although homology modeling techniques are widely used to produce low-resolution models, refining these models to high resolution has proven difficult. With long enough simulations and sufficiently accurate force fields, molecular dynamics (MD) simulations should in principle allow such refinement, but efforts to refine homology models using MD have for the most part yielded disappointing results. It has thus far been unclear whether MD-based refinement is limited primarily by accessible simulation timescales, force field accuracy, or both. Here, we examine MD as a technique for homology model refinement using all-atom simulations, each at least 100 µs long-more than 100 times longer than previous refinement simulations-and a physics-based force field that was recently shown to successfully fold a structurally diverse set of fast-folding proteins. In MD simulations of 24 proteins chosen from the refinement category of recent Critical Assessment of Structure Prediction (CASP) experiments, we find that in most cases, simulations initiated from homology models drift away from the native structure. Comparison with simulations initiated from the native structure suggests that force field accuracy is the primary factor limiting MD-based refinement. This problem can be mitigated to some extent by restricting sampling to the neighborhood of the initial model, leading to structural improvement that, while limited, is roughly comparable to the leading alternative methods.


Assuntos
Modelos Moleculares , Simulação de Dinâmica Molecular , Proteínas/química , Homologia Estrutural de Proteína , Biologia Computacional/métodos , Conformação Proteica , Dobramento de Proteína
18.
PLoS One ; 7(2): e32131, 2012.
Artigo em Inglês | MEDLINE | ID: mdl-22384157

RESUMO

Molecular dynamics simulations provide a vehicle for capturing the structures, motions, and interactions of biological macromolecules in full atomic detail. The accuracy of such simulations, however, is critically dependent on the force field--the mathematical model used to approximate the atomic-level forces acting on the simulated molecular system. Here we present a systematic and extensive evaluation of eight different protein force fields based on comparisons of experimental data with molecular dynamics simulations that reach a previously inaccessible timescale. First, through extensive comparisons with experimental NMR data, we examined the force fields' abilities to describe the structure and fluctuations of folded proteins. Second, we quantified potential biases towards different secondary structure types by comparing experimental and simulation data for small peptides that preferentially populate either helical or sheet-like structures. Third, we tested the force fields' abilities to fold two small proteins--one α-helical, the other with ß-sheet structure. The results suggest that force fields have improved over time, and that the most recent versions, while not perfect, provide an accurate description of many structural and dynamical properties of proteins.


Assuntos
Proteínas/química , Biologia Computacional/métodos , Simulação por Computador , Humanos , Espectroscopia de Ressonância Magnética/métodos , Modelos Moleculares , Conformação Molecular , Simulação de Dinâmica Molecular , Peptídeos/química , Dobramento de Proteína , Estrutura Secundária de Proteína , Estrutura Terciária de Proteína , Temperatura , Ubiquitina/química
19.
J Am Chem Soc ; 133(24): 9181-3, 2011 Jun 22.
Artigo em Inglês | MEDLINE | ID: mdl-21545110

RESUMO

Although the thermodynamic principles that control the binding of drug molecules to their protein targets are well understood, detailed experimental characterization of the process by which such binding occurs has proven challenging. We conducted relatively long, unguided molecular dynamics simulations in which a ligand (the cancer drug dasatinib or the kinase inhibitor PP1) was initially placed at a random location within a box that also contained a protein (Src kinase) to which that ligand was known to bind. In several of these simulations, the ligand correctly identified its target binding site, forming a complex virtually identical to the crystallographically determined bound structure. The simulated trajectories provide a continuous, atomic-level view of the entire binding process, revealing persistent and noteworthy intermediate conformations and shedding light on the role of water molecules. The technique we employed, which does not assume any prior knowledge of the binding site's location, may prove particularly useful in the development of allosteric inhibitors that target previously undiscovered binding sites.


Assuntos
Simulação de Dinâmica Molecular , Pirazóis/metabolismo , Pirimidinas/metabolismo , Tiazóis/metabolismo , Sítios de Ligação , Dasatinibe , Ligação Proteica , Conformação Proteica , Pirazóis/farmacologia , Pirimidinas/farmacologia , Quinases da Família src/antagonistas & inibidores , Quinases da Família src/química , Quinases da Família src/metabolismo
20.
Science ; 330(6002): 341-6, 2010 Oct 15.
Artigo em Inglês | MEDLINE | ID: mdl-20947758

RESUMO

Molecular dynamics (MD) simulations are widely used to study protein motions at an atomic level of detail, but they have been limited to time scales shorter than those of many biologically critical conformational changes. We examined two fundamental processes in protein dynamics--protein folding and conformational change within the folded state--by means of extremely long all-atom MD simulations conducted on a special-purpose machine. Equilibrium simulations of a WW protein domain captured multiple folding and unfolding events that consistently follow a well-defined folding pathway; separate simulations of the protein's constituent substructures shed light on possible determinants of this pathway. A 1-millisecond simulation of the folded protein BPTI reveals a small number of structurally distinct conformational states whose reversible interconversion is slower than local relaxations within those states by a factor of more than 1000.


Assuntos
Aprotinina/química , Simulação de Dinâmica Molecular , Conformação Proteica , Dobramento de Proteína , Proteínas/química , Substituição de Aminoácidos , Biologia Computacional , Computadores , Cinética , Proteínas dos Microfilamentos/química , Modelos Moleculares , Proteínas Mutantes/química , Estrutura Terciária de Proteína , Solventes , Termodinâmica
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