75 research outputs found
Smyd2 conformational changes in response to p53 binding: role of the C-terminal domain.
Smyd2 lysine methyltransferase regulates monomethylation of histone and nonhistone lysine residues using S-adenosylmethionine cofactor as the methyl donor. The nonhistone interactors include several tumorigenic targets, including p53. Understanding this interaction would allow the structural principles that underpin Smyd2-mediated p53 methylation to be elucidated. Here, we performed μ-second molecular dynamics (MD) simulations on binary Smyd2-cofactor and ternary Smyd2-cofactor-p53 peptide complexes. We considered both unmethylated and monomethylated p53 peptides (at Lys370 and Lys372). The results indicate that (a) the degree of conformational freedom of the C-terminal domain of Smyd2 is restricted by the presence of the p53 peptide substrate, (b) the Smyd2 C-terminal domain shows distinct dynamic properties when interacting with unmethylated and methylated p53 peptides, and (c) Lys372 methylation confines the p53 peptide conformation, with detectable influence on Lys370 accessibility to the cofactor. These MD results are therefore of relevance for studying the biology of p53 in cancer progression
Assessment of Multi-Scale Approaches for ComputingUV–Vis Spectra in Condensed Phases: Toward an Effective yetReliable Integration of Variational and Perturbative QM/MM Approaches
Computational simulation of UV/vis spectra in condensed phases can be performed starting from converged molecular dynamics (MD) simulations and then performing quantum mechanical/molecular mechanical (QM/MM) computations for a statistically significant number of snapshots. However, the need of variational solutions (e.g., ONIOM/EE) for a huge number of snapshots makes unpractical the use of state-of-the-art QM Hamiltonians. On the other hand, the effectivity of perturbative approaches (e.g., perturbed matrix method, PMM) comes at the price of poor convergence for configurations strongly different from the reference one. In this paper we introduce an integrated strategy based on a cluster analysis of the MD snapshots. Next, a representative configuration for each cluster is treated at the ONIOM/EE level, whereas local fluctuations within each cluster are described at the PMM level. Some representative systems (uracil in dimethylformamide and in water and tyrosine zwitterion in water) are analyzed to show t..
Electrostatic and Structural Bases of Fe2+ Translocation through Ferritin Channels
Ferritin molecular cages are marvelous 24-mer supramolecular architectures that enable massive iron storage (>2000 iron atoms) within their inner cavity. This cavity is connected to the outer environment by two channels at C3 and C4 symmetry axes of the assembly. Ferritins can also be exploited as carriers for in vivo imaging and therapeutic applications, owing to their capability to effectively protect synthetic non-endogenous agents within the cage cavity and deliver them to targeted tissue cells without stimulating adverse immune responses. Recently, X-ray crystal structures of Fe(2+)-loaded ferritins provided important information on the pathways followed by iron ions toward the ferritin cavity and the catalytic centers within the protein. However, the specific mechanisms enabling Fe(2+) uptake through wild-type and mutant ferritin channels is largely unknown. To shed light on this question, we report extensive molecular dynamics simulations, site-directed mutagenesis, and kinetic measurements that characterize the transport properties and translocation mechanism of Fe(2+) through the two ferritin channels, using the wild-type bullfrog Rana catesbeiana H' protein and some of its variants as case studies. We describe the structural features that determine Fe(2+) translocation with atomistic detail, and we propose a putative mechanism for Fe(2+) transport through the channel at the C3 symmetry axis, which is the only iron-permeable channel in vertebrate ferritins. Our findings have important implications for understanding how ion permeation occurs, and further how it may be controlled via purposely engineered channels for novel biomedical applications based on ferritin
Computational Study of Helicase from SARS-CoV-2 in RNA-Free and Engaged Form
Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is the causative agent of the pandemic that broke out in 2020 and continues to be the cause of massive global upheaval. Coronaviruses are positive-strand RNA viruses with a genome of ~30 kb. The genome is replicated and transcribed by RNA-dependent RNA polymerase together with accessory factors. One of the latter is the protein helicase (NSP13), which is essential for viral replication. The recently solved helicase structure revealed a tertiary structure composed of five domains. Here, we investigated NSP13 from a structural point of view, comparing its RNA-free form with the RNA-engaged form by using atomistic molecular dynamics (MD) simulations at the microsecond timescale. Structural analyses revealed conformational changes that provide insights into the contribution of the different domains, identifying the residues responsible for domain–domain interactions in both observed forms. The RNA-free system appears to be more flexible than the RNA-engaged form. This result underlies the stabilizing role of the nucleic acid and the functional core role of these domains
Effective yet Reliable Computation of EPR Spectra in Solution by a QM/MM Approach: Interplay between Electrostatics and Non-electrostatic Effects
In this paper, we have extended to the calculation of hyperfine coupling constants, the model recently proposed by some of the present authors
[Giovannini et al., J. Chem. Theory Comput. 13, 4854\u20134870 (2017)] to include Pauli repulsion and dispersion effects in Quantum Mechanical/
Molecular Mechanics (QM/MM) approaches. The peculiarity of the proposed approach stands in the fact that repulsion/dispersion
contributions are explicitly introduced in the QM Hamiltonian. Therefore, such terms not only enter the evaluation of energetic properties
but also propagate to molecular properties and spectra. A novel parametrization of the electrostatic fluctuating charge force field has
been developed, thus allowing a quantitative reproduction of reference QM interaction energies. Such a parametrization has been then tested
against the prediction of EPR parameters of prototypical nitroxide radicals in aqueous solutions
Non-linear dimensionality reduction on extracellular waveforms reveals cell type diversity in premotor cortex
Cortical circuits are thought to contain a large number of cell types that coordinate to produce behavior. Current in vivo methods rely on clustering of specified features of extracellular waveforms to identify putative cell types, but these capture only a small amount of variation. Here, we develop a new method (WaveMAP) that combines non-linear dimensionality reduction with graph clustering to identify putative cell types. We apply WaveMAP to extracellular waveforms recorded from dorsal premotor cortex of macaque monkeys performing a decision-making task. Using WaveMAP, we robustly establish eight waveform clusters and show that these clusters recapitulate previously identified narrow- and broad-spiking types while revealing previously unknown diversity within these subtypes. The eight clusters exhibited distinct laminar distributions, characteristic firing rate patterns, and decision-related dynamics. Such insights were weaker when using feature-based approaches. WaveMAP therefore provides a more nuanced understanding of the dynamics of cell types in cortical circuits.https://elifesciences.org/articles/67490Published versio
Hydration pattern of A4T4 and T4A4 DNA: a molecular dynamics study
Hydration pattern and energetics of ‘A-tract’ containing duplexes have been studied using molecular dynamics on 12-mer self-complementary sequences 5′-d(GCA4T4GC)-3′ and 5′-d(CGT4A4CG)-3′. The structural features for the simulated duplexes showed correlation with the corresponding experimental structures. Analysis of the hydration pattern confirmed that water network around the simulated duplexes is more conformation specific rather than sequence specific. The calculated heat capacity change upon duplex formation showed that the process is entropically driven for both the sequences. Furthermore, the theoretical free energy estimates calculated using MMPBSA approach showed a higher net electrostatic contribution for A4T4 duplex formation than for T4A4, however, energetically both the duplexes are observed to be equally stable
Pathways and Barriers for Ion Translocation through the 5-HT3A Receptor Channel
Pentameric ligand gated ion channels (pLGICs) are ionotropic receptors that mediate fast intercellular communications at synaptic level and include either cation selective (e.g., nAChR and 5-HT3) or anion selective (e.g., GlyR, GABAA and GluCl) membrane channels. Among others, 5-HT3 is one of the most studied members, since its first cloning back in 1991, and a large number of studies have successfully pinpointed protein residues critical for its activation and channel gating. In addition, 5-HT3 is also the target of a few pharmacological treatments due to the demonstrated benefits of its modulation in clinical trials. Nonetheless, a detailed molecular analysis of important protein features, such as the origin of its ion selectivity and the rather low conductance as compared to other channel homologues, has been unfeasible until the recent crystallization of the mouse 5-HT3A receptor. Here, we present extended molecular dynamics simulations and free energy calculations of the whole 5-HT3A protein with the aim of better understanding its ion transport properties, such as the pathways for ion permeation into the receptor body and the complex nature of the selectivity filter. Our investigation unravels previously unpredicted structural features of the 5-HT3A receptor, such as the existence of alternative intersubunit pathways for ion translocation at the interface between the extracellular and the transmembrane domains, in addition to the one along the channel main axis. Moreover, our study offers a molecular interpretation of the role played by an arginine triplet located in the intracellular domain on determining the characteristic low conductance of the 5-HT3A receptor, as evidenced in previous experiments. In view of these results, possible implications on other members of the superfamily are suggested
Conformational Dynamics of Lysine Methyltransferase Smyd2. Insights into the Different Substrate Crevice Characteristics of Smyd2 and Smyd3
Smyd2, the SET and MYND domain containing
protein lysine methyltransferase,
targets histone and nonhistone substrates. Methylation of nonhistone
substrates has direct implications in cancer development and progression.
Dynamic regulation of Smyd2 activity and the structural basis of broad
substrate specificity still remain elusive. Herein, we report on extensive
molecular dynamics simulations on a full length Smyd2 in the presence
and absence of AdoMet cofactor (covering together 1.3 μs of
sampling), and the accompanying conformational transitions. Additionally,
dynamics of the C-terminal domain (CTD) and structural features of
substrate crevices of Smyd2 and Smyd3 are compared. The CTD of Smyd2
exhibits conformational flexibility in both states. In the holo form,
however, it undergoes larger hinge motions resulting in more opened
configurations than the apo form, which is confined around the partially
open starting X-ray configuration. AdoMet binding triggers increased
elasticity of the CTD leading Smyd2 to adopt fully opened configurations,
which completely exposes the substrate binding crevice. These long-range
concerted motions highlight Smyd2’s ability to target substrates
of varying sizes. Substrate crevices of Smyd2 and Smyd3 show distinct
features in terms of spatial, hydration, and electrostatic properties
that emphasize their characteristic modes of substrates interaction
and entry pathways for inhibitor binding. On the whole, our study
shows how the elasticity and hinge motion of the CTD regulate its
functional role and underpin the basis of broad substrate specificity
of Smyd2. We also highlight the specific structural principles that
guide substrate and inhibitor binding to Smyd2 and Smyd3
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