196,011 research outputs found
Microtubule assembly governed by tubulin allosteric gain in flexibility and lattice induced fit.
Get PDFMicrotubules (MTs) are key components of the cytoskeleton and play a central role in cell division and development. MT assembly is known to be associated with a structural change in αβ-tubulin dimers from kinked to straight conformations. How GTP binding renders individual dimers polymerization-competent, however, is still unclear. Here, we have characterized the conformational dynamics and energetics of unassembled tubulin using atomistic molecular dynamics and free energy calculations. Contradictory to existing allosteric and lattice models, we find that GTP-tubulin favors a broad range of almost isoenergetic curvatures, whereas GDP-tubulin has a much lower bending flexibility. Moreover, irrespective of the bound nucleotide and curvature, two conformational states exist differing in location of the anchor point connecting the monomers that affects tubulin bending, with one state being strongly favored in solution. Our findings suggest a new combined model in which MTs incorporate and stabilize flexible GTP-dimers with a specific anchor point state
Bending-torsional elasticity and energetics of the plus-end microtubule tip
Get PDFMicrotubules (MTs), mesoscopic cellular filaments, grow primarily by the additionof GTP-bound tubulin dimers at their dynamic flaring plus-end tips. They operateas chemomechanical energy transducers with stochastic transitions to an astoundingshortening motion upon hydrolyzing GTP to GDP. Time-resolved dynamics of theMT tip—a key determinant of this behavior—as a function of nucleotide state, internallattice strain, and stabilizing lateral interactions have not been fully understood. Herewe use atomistic simulations to study the spontaneous relaxation of complete GTP-MTand GDP-MT tip models from unfavorable straight to relaxed splayed conformationsand to comprehensively characterize the elasticity of MT tips. Our simulations revealthe dominance of viscoelastic dynamics of MT protofilaments during the relaxationprocess, driven by the stored bending-torsional strain and counterbalanced by theinterprotofilament interactions. We show that the posthydrolysis MT tip is exposedto higher activation energy barriers for straight lattice formation, which translates intoits inability to elongate. Our study provides an information-driven Brownian ratchetmechanism for the elastic energy conversion and release by MT tips and offers insightsinto the mechanoenzymatics of MTs
Single-particle Cryo-EM and molecular dynamics simulations: A perfect match
Get PDFFunder:
http://dx.doi.org/10.13039/501100001659
Deutsche Forschungsgemeinschaf
Choice of fluorophore affects dynamic DNA nanostructures
Get PDFThe ability to dynamically remodel DNA origami structures or functional nanodevices is highly desired in the field of DNA nanotechnology. Concomitantly, the use of fluorophores to track and validate the dynamics of such DNA-based architectures is commonplace and often unavoidable. It is therefore crucial to be aware of the side effects of popular fluorophores, which are often exchanged without considering the potential impact on the system. Here, we show that the choice of fluorophore can strongly affect the reconfiguration of DNA nanostructures. To this end, we encapsulate a triple-stranded DNA (tsDNA) into water-in-oil compartments and functionalize their periphery with a single-stranded DNA handle (ssDNA). Thus, the tsDNA can bind and unbind from the periphery by reversible opening of the triplex and subsequent strand displacement. Using a combination of experiments, molecular dynamics (MD) simulations, and reaction-diffusion modeling, we demonstrate for twelve different fluorophore combinations that it is possible to alter or even inhibit the DNA nanostructure formation – without changing the DNA sequence. Besides its immediate importance for the design of pH-responsive switches and fluorophore labelling, our work presents a strategy to precisely tune the energy landscape of dynamic DNA nanodevices
Microtubule dynamics are defined by conformations and stability of clustered protofilaments
Get PDFMicrotubules are dynamic cytoskeletal polymers that add and lose tubulin dimers at their ends. Microtubule growth, shortening, and transitions between them are linked to GTP hydrolysis. Recent evidence suggests that flexible tubulin protofilaments at microtubule ends adopt a variety of shapes, complicating structural analysis using conventional techniques. Therefore, the link between GTP hydrolysis, protofilament structure and microtubule polymerization state is poorly understood. Here, we investigate the conformational dynamics of microtubule ends using coarse-grained modeling supported by atomistic simulations and cryoelectron tomography. We show that individual bent protofilaments organize in clusters, transient precursors to the straight microtubule lattice, with GTP-bound ends showing elevated and more persistent cluster formation. Differences in the mechanical properties of GTP- and GDP-protofilaments result in differences in intracluster tension, determining both clustering propensity and protofilament length. We propose that conformational selection at microtubule ends favors long-lived clusters of short GTP-protofilaments that are more prone to forming a straight microtubule lattice and accommodating new tubulin dimers. Conversely, microtubule ends trapped in states with unevenly long and stiff GDP-protofilaments are more prone to shortening. We conclude that protofilament clustering is the key phenomenon that links the hydrolysis state of single tubulins to the polymerization state of the entire microtubule
Microtubule dynamics are defined by conformations and stability of clustered protofilaments
No full textMicrotubules are dynamic cytoskeletal polymers that add and lose tubulin dimers at their ends. Microtubule growth, shortening and transitions between them are linked to GTP hydrolysis. Recent evidence suggests that flexible tubulin protofilaments at microtubule ends adopt a variety of shapes, complicating structural analysis using conventional techniques. Therefore, the link between GTP hydrolysis, protofilament structure and microtubule polymerization state is poorly understood. Here, we investigate the conformational dynamics of microtubule ends using coarse-grained modeling supported by atomistic simulations and cryo-electron tomography. We show that individual bent protofilaments organize in clusters, transient precursors to a straight
microtubule lattice, with GTP-bound ends showing elevated and more persistent cluster formation. Differences in the mechanical properties of GTP- and GDP-protofilaments result in differences in intra-cluster tension, determining both clustering propensity and protofilament length. We propose that conformational selection at microtubule ends favors long-lived clusters of short GTP-protofilaments that are more prone to form a straight microtubule lattice and accommodate new tubulin dimers. Conversely, microtubule ends trapped in states with
unevenly long and stiff GDP-protofilaments are more prone to shortening. We conclude that protofilament clustering is a key phenomenon that links the hydrolysis state of single tubulins to the polymerization state of the entire microtubule
SESCA: Predicting circular dichroism spectra from protein molecular structures
Get PDFCircular dichroism spectroscopy is a highly sensitive, but low-resolution technique to study the structure of proteins. Combed with molecular modelling and other complementary techniques, CD spectroscopy can also provide essential information at higher resolution. To this aim, we introduce a new computational method to calculate the electronic circular dichroism spectra of proteins from a three dimensional-model structure or structural ensemble. The method determines the CD spectrum from the average secondary structure composition of the protein using a pre-calculated set of basis spectra. We derived several basis spectrum sets obtained from the experimental CD spectra and secondary structure information of 71 reference proteins and tested the prediction accuracy of these basis spectrum sets through cross-validation. Furthermore, we investigated how prediction accuracy is affected by contributions from amino acid side chain groups and protein flexibility, potential experimental errors of the reference protein spectra, as well as the choice of the secondary structure classification algorithm and the number of basis spectra. We compared the predictive power of our method to previous spectrum prediction algorithms — such as DichroCalc and PDB2CD — and found that SESCA predicts the CD spectra with up to 50% smaller deviation. Our results indicate that SESCA basis sets are robust to experimental error in the reference spectra, and the choice of the secondary structure classification algorithm. For over 80% of the globular reference proteins, SESCA basis sets could accurately predict the experimental spectrum solely from their secondary structure composition. To improve SESCA predictions for the remaining proteins, we applied corrections to account for intensity normalization, contributions from the amino side chains, and conformational flexibility. For globular proteins only intensity scaling improved the prediction accuracy significantly, but our models indicate that side chain contributions and structural flexibility are pivotal for the prediction of shorter peptides and intrinsically disordered proteins
Automated cryo-EM structure refinement using correlation-driven molecular dynamics.
Get PDFWe present a correlation-driven molecular dynamics (CDMD) method for automated refinement of atomistic models into cryo-electron microscopy (cryo-EM) maps at resolutions ranging from near-atomic to subnanometer. It utilizes a chemically accurate force field and thermodynamic sampling to improve the real-space correlation between the modeled structure and the cryo-EM map. Our framework employs a gradual increase in resolution and map-model agreement as well as simulated annealing, and allows fully automated refinement without manual intervention or any additional rotamer- and backbone-specific restraints. Using multiple challenging systems covering a wide range of map resolutions, system sizes, starting model geometries and distances from the target state, we assess the quality of generated models in terms of both model accuracy and potential of overfitting. To provide an objective comparison, we apply several well-established methods across all examples and demonstrate that CDMD performs best in most cases
Dr. Duane M. Jackson, Morehouse College, July 2011
No full textThis video is a conversation with Dr. Duane M. Jackson. Dr. Jackson talks about his paper, "Recall and the Serial Position Effect: The Role of Primacy and Recency on Accounting Students' Performance." Jackie Daniel, AUC Woodruff Library, is the interviewer
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