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Find video protocols related to scientific articles indexed in Pubmed.
The yeast model suggests the use of short peptides derived from mt LeuRS for the therapy of diseases due to mutations in several mt tRNAs.
Biochim. Biophys. Acta
PUBLISHED: 07-02-2014
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We have previously established a yeast model of mitochondrial (mt) diseases. We showed that defective respiratory phenotypes due to point-mutations in mt tRNA(Leu(UUR)), tRNA(Ile) and tRNA(Val) could be relieved by overexpression of both cognate and non-cognate nuclearly encoded mt aminoacyl-tRNA synthetases (aaRS) LeuRS, IleRS and ValRS. More recently, we showed that the isolated carboxy-terminal domain (Cterm) of yeast mt LeuRS, and even short peptides derived from the human Cterm, have the same suppressing abilities as the whole enzymes. In this work, we extend these results by investigating the activity of a number of mt aaRS from either class I or II towards a panel of mt tRNAs. The Cterm of both human and yeast mt LeuRS has the same spectrum of activity as mt aaRS belonging to class I and subclass a, which is the most extensive among the whole enzymes. Yeast Cterm is demonstrated to be endowed with mt targeting activity. Importantly, peptide fragments ?30_31 and ?32_33, derived from the human Cterm, have even higher efficiency as well as wider spectrum of activity, thus opening new avenues for therapeutic intervention. Bind-shifting experiments show that the ?30_31 peptide directly interacts with human mt tRNA(Leu(UUR)) and tRNA(Ile), suggesting that the rescuing activity of isolated peptide fragments is mediated by a chaperone-like mechanism. Wide-range suppression appears to be idiosyncratic of LeuRS and its fragments, since it is not shared by Cterminal regions derived from human mt IleRS or ValRS, which are expected to have very different structures and interactions with tRNAs.
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Strain-specific nuclear genetic background differentially affects mitochondria-related phenotypes in Saccharomyces cerevisiae.
Microbiologyopen
PUBLISHED: 01-09-2014
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In the course of our studies on mitochondrial defects, we have observed important phenotypic variations in Saccharomyces cerevisiae strains suggesting that a better characterization of the genetic variability will be essential to define the relationship between the mitochondrial efficiency and the presence of different nuclear backgrounds. In this manuscript, we have extended the study of such relations by comparing phenotypic assays related to mitochondrial functions of three wild-type laboratory strains. In addition to the phenotypic variability among the wild-type strains, important differences have been observed among strains bearing identical mitochondrial tRNA mutations that could be related only to the different nuclear background of the cells. Results showed that strains exhibited an intrinsic variability in the severity of the effects of the mitochondrial mutations and that specific strains might be used preferentially to evaluate the phenotypic effect of mitochondrial mutations on carbon metabolism, stress responses, and mitochondrial DNA stability. In particular, while W303-1B and MCC123 strains should be used to study the effect of severe mitochondrial tRNA mutations, D273-10B/A1 strain is rather suitable for studying the effects of milder mutations.
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Physical properties of mixed bilayers containing lamellar and nonlamellar lipids: insights from coarse-grain molecular dynamics simulations.
Faraday Discuss.
PUBLISHED: 06-29-2013
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A recently developed coarse-grain model is applied to simulate hydrated membranes containing the lamellar lipid DOPC and the nonlamellar lipid DOPE. In a first series of simulations, DOPC-water and DOPE-water systems are shown to form respectively bilayers and inverse hexagonal phases, in agreement with the well-known behaviour observed experimentally. A second set of calculations is then run to investigate several fundamental physical features of mixed DOPC-DOPE bilayers at different relative compositions. In particular, a quantitative characterisation is obtained of the internal distributions (profiles) of lateral pressure and electrical potential. These two properties, very difficult to measure experimentally, are thought to underpin many key membrane phenomena, including nonspecific lipid-mediated mechanisms of protein regulation. The molecular origin of the distributions, and their dependence on changes in the DOPC: DOPE ratio, are explained through an analysis of separate contributions from individual interaction types and molecular groups.
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Analyzing the suppression of respiratory defects in the yeast model of human mitochondrial tRNA diseases.
Gene
PUBLISHED: 02-01-2013
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The respiratory defects associated with mutations in human mitochondrial tRNA genes can be mimicked in yeast, which is the only organism easily amenable to mitochondrial transformation. This approach has shown that overexpression of several nuclear genes coding for factors involved in mitochondrial protein synthesis can alleviate the respiratory defects both in yeast and in human cells. The present paper analyzes in detail the effects of overexpressed yeast and human mitochondrial translation elongation factors EF-Tu. We studied the suppressing activity versus the function in mt translation of mutated versions of this factor and we obtained indications on the mechanism of suppression. Moreover from a more extended search for suppressor genes we isolated factors which might be active in mitochondrial biogenesis. Results indicate that the multiplicity of mitochondrial factors as well as their high variability of expression levels can account for the variable severity of mitochondrial diseases and might suggest possible therapeutic approaches.
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The ELBA force field for coarse-grain modeling of lipid membranes.
PLoS ONE
PUBLISHED: 08-02-2011
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A new coarse-grain model for molecular dynamics simulation of lipid membranes is presented. Following a simple and conventional approach, lipid molecules are modeled by spherical sites, each representing a group of several atoms. In contrast to common coarse-grain methods, two original (interdependent) features are here adopted. First, the main electrostatics are modeled explicitly by charges and dipoles, which interact realistically through a relative dielectric constant of unity (?(r) = 1). Second, water molecules are represented individually through a new parametrization of the simple Stockmayer potential for polar fluids; each water molecule is therefore described by a single spherical site embedded with a point dipole. The force field is shown to accurately reproduce the main physical properties of single-species phospholipid bilayers comprising dioleoylphosphatidylcholine (DOPC) and dioleoylphosphatidylethanolamine (DOPE) in the liquid crystal phase, as well as distearoylphosphatidylcholine (DSPC) in the liquid crystal and gel phases. Insights are presented into fundamental properties and phenomena that can be difficult or impossible to study with alternative computational or experimental methods. For example, we investigate the internal pressure distribution, dipole potential, lipid diffusion, and spontaneous self-assembly. Simulations lasting up to 1.5 microseconds were conducted for systems of different sizes (128, 512 and 1058 lipids); this also allowed us to identify size-dependent artifacts that are expected to affect membrane simulations in general. Future extensions and applications are discussed, particularly in relation to the methodologys inherent multiscale capabilities.
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Dual-resolution molecular dynamics simulation of antimicrobials in biomembranes.
J R Soc Interface
PUBLISHED: 12-03-2010
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Triclocarban and triclosan, two potent antibacterial molecules present in many consumer products, have been subject to growing debate on a number of issues, particularly in relation to their possible role in causing microbial resistance. In this computational study, we present molecular-level insights into the interaction between these antimicrobial agents and hydrated phospholipid bilayers (taken as a simple model for the cell membrane). Simulations are conducted by a novel dual-resolution molecular dynamics approach which combines accuracy with efficiency: the antimicrobials, modelled atomistically, are mixed with simplified (coarse-grain) models of lipids and water. A first set of calculations is run to study the antimicrobials transfer free energies and orientations as a function of depth inside the membrane. Both molecules are predicted to preferentially accumulate in the lipid headgroup-glycerol region; this finding, which reproduces corresponding experimental data, is also discussed in terms of a general relation between solute partitioning and the intramembrane distribution of pressure. A second set of runs involves membranes incorporated with different molar concentrations of antimicrobial molecules (up to one antimicrobial per two lipids). We study the effects induced on fundamental membrane properties, such as the electron density, lateral pressure and electrical potential profiles. In particular, the analysis of the spontaneous curvature indicates that increasing antimicrobial concentrations promote a destabilizing tendency towards non-bilayer phases, as observed experimentally. The antimicrobials influence on the self-assembly process is also investigated. The significance of our results in the context of current theories of antimicrobial action is discussed.
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Anisotropic elastic network modeling of entire microtubules.
Biophys. J.
PUBLISHED: 06-19-2010
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Microtubules are supramolecular structures that make up the cytoskeleton and strongly affect the mechanical properties of the cell. Within the cytoskeleton filaments, the microtubule (MT) exhibits by far the highest bending stiffness. Bending stiffness depends on the mechanical properties and intermolecular interactions of the tubulin dimers (the MT building blocks). Computational molecular modeling has the potential for obtaining quantitative insights into this area. However, to our knowledge, standard molecular modeling techniques, such as molecular dynamics (MD) and normal mode analysis (NMA), are not yet able to simulate large molecular structures like the MTs; in fact, their possibilities are normally limited to much smaller protein complexes. In this work, we developed a multiscale approach by merging the modeling contribution from MD and NMA. In particular, MD simulations were used to refine the molecular conformation and arrangement of the tubulin dimers inside the MT lattice. Subsequently, NMA was used to investigate the vibrational properties of MTs modeled as an elastic network. The coarse-grain model here developed can describe systems of hundreds of interacting tubulin monomers (corresponding to up to 1,000,000 atoms). In particular, we were able to simulate coarse-grain models of entire MTs, with lengths up to 350 nm. A quantitative mechanical investigation was performed; from the bending and stretching modes, we estimated MT macroscopic properties such as bending stiffness, Young modulus, and persistence length, thus allowing a direct comparison with experimental data.
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Coarse-grain modelling of DMPC and DOPC lipid bilayers.
J Phys Condens Matter
PUBLISHED: 03-09-2010
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Our recently developed coarse-grain model for dimyristoylphosphatidylcholine (DMPC) has been improved and extended to dioleylphosphatidylcholine (DOPC), a more typical constituent of real biological membranes. Single-component DMPC and DOPC bilayers have been simulated using microsecond-long molecular dynamics. We investigated properties that are difficult or impossible to access experimentally, such as the pressure distribution, the spontaneous curvature and the diffusion pattern of individual lipid molecules. Moreover, we studied the dipole potential, a basic physical feature of paramount biological importance that cannot be currently modelled by other coarse-grain approaches. In fact, a complete representation of the system electrostatics and a realistic description of the water component make our method unique amongst the existing coarse-grain membrane models. The spontaneous permeation of water, a phenomenon out of reach of standard atomistic models, was also observed and quantified; this was possible thanks to the efficiency of our model, which is about two orders of magnitude less computationally expensive than atomic-level counterparts. Results are generally in good agreement with the literature data. Further model extensions and future applications are proposed.
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Permeability of small molecules through a lipid bilayer: a multiscale simulation study.
J Phys Chem B
PUBLISHED: 08-12-2009
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The transmembrane permeation of eight small (molecular weight <100) organic molecules across a phospholipid bilayer is investigated by multiscale molecular dynamics simulation. The bilayer and hydrating water are represented by simplified, efficient coarse-grain models, whereas the permeating molecules are described by a standard atomic-level force-field. Permeability properties are obtained through a refined version of the z-constraint algorithm. By constraining each permeant at selected depths inside the bilayer, we have sampled free energy differences and diffusion coefficients across the membrane. These data have been combined, according to the inhomogeneous solubility-diffusion model, to yield the permeability coefficients. The results are generally consistent with previous atomic-level calculations and available experimental data. Computationally, our multiscale approach proves 2 orders of magnitude faster than traditional atomic-level methods.
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Acceleration of coarse grain molecular dynamics on GPU architectures.
J Comput Chem
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Coarse grain (CG) molecular models have been proposed to simulate complex systems with lower computational overheads and longer timescales with respect to atomistic level models. However, their acceleration on parallel architectures such as graphic processing units (GPUs) presents original challenges that must be carefully evaluated. The objective of this work is to characterize the impact of CG model features on parallel simulation performance. To achieve this, we implemented a GPU-accelerated version of a CG molecular dynamics simulator, to which we applied specific optimizations for CG models, such as dedicated data structures to handle different bead type interactions, obtaining a maximum speed-up of 14 on the NVIDIA GTX480 GPU with Fermi architecture. We provide a complete characterization and evaluation of algorithmic and simulated system features of CG models impacting the achievable speed-up and accuracy of results, using three different GPU architectures as case studies.
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What is Visualize?

JoVE Visualize is a tool created to match the last 5 years of PubMed publications to methods in JoVE's video library.

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We use abstracts found on PubMed and match them to JoVE videos to create a list of 10 to 30 related methods videos.

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In developing our video relationships, we compare around 5 million PubMed articles to our library of over 4,500 methods videos. In some cases the language used in the PubMed abstracts makes matching that content to a JoVE video difficult. In other cases, there happens not to be any content in our video library that is relevant to the topic of a given abstract. In these cases, our algorithms are trying their best to display videos with relevant content, which can sometimes result in matched videos with only a slight relation.