Increasing cleavage specificity and activity of restriction endonuclease KpnI

Restriction enzyme KpnI is a HNH superfamily endonuclease requiring divalent metal ions for DNA cleavage but not for binding. The active site of KpnI can accommodate metal ions of different atomic radii for DNA cleavage. Although Mg2+ ion higher than 500 mu M mediates promiscuous activity, Ca2+ suppresses the promiscuity and induces high cleavage fidelity. Here, we report that a conservative mutation of the metal-coordinating residue D148 to Glu results in the elimination of the Ca2+-mediated cleavage but imparting high cleavage fidelity with Mg2+. High cleavage fidelity of the mutant D148E is achieved through better discrimination of the target site at the binding and cleavage steps. Biochemical experiments and molecular dynamics simulations suggest that the mutation inhibits Ca2+-mediated cleavage activity by altering the geometry of the Ca2+-bound HNH active site. Although the D148E mutant reduces the specific activity of the enzyme, we identified a suppressor mutation that increases the turnover rate to restore the specific activity of the high fidelity mutant to the wild-type level. Our results show that active site plasticity in coordinating different metal ions is related to KpnI promiscuous activity, and tinkering the metal ion coordination is a plausible way to reduce promiscuous activity of metalloenzymes.

Electronic band Structure and Photoemission Spectra of Graphene on Silicon Substrate

Synergizing graphene on silicon based nanostructures is pivotal in advancing nano-electronic device technology. A combination of molecular dynamics and density functional theory has been used to predict the electronic energy band structure and photo-emission spectrum for graphene-Si system with silicon as a substrate for graphene. The equilibrium geometry of the system after energy minimization is obtained from molecular dynamics simulations. For the stable geometry obtained, density functional theory calculations are employed to determine the energy band structure and dielectric constant of the system. Further the work function of the system which is a direct consequence of photoemission spectrum is calculated from the energy band structure using random phase approximations.

Defect induced plasticity and failure mechanism of boron nitride nanotubes under tension

The effects of Stone-Wales (SW) and vacancy defects on the failure behavior of boron nitride nanotubes (BNNTs) under tension are investigated using molecular dynamics simulations. The Tersoff-Brenner potential is used to model the atomic interaction and the temperature is maintained close to 300 K. The effect of a SW defect is studied by determining the failure strength and failure mechanism of nanotubes with different radii. In the case of a vacancy defect, the effect of an N-vacancy and a B-vacancy is studied separately. Nanotubes with different chiralities but similar diameter is considered first to evaluate the chirality dependence. The variation of failure strength with the radius is then studied by considering nanotubes of different diameters but same chirality. It is observed that the armchair BNNTs are extremely sensitive to defects, whereas the zigzag configurations are the least sensitive. In the case of pristine BNNTs, both armchair and zigzag nanotubes undergo brittle failure, whereas in the case of defective BNNTs, only the zigzag ones undergo brittle failure. An interesting defect induced plastic behavior is observed in defective armchair BNNTs. For this nanotube, the presence of a defect triggers mechanical relaxation by bond breaking along the closest zigzag helical path, with the defect as the nucleus. This mechanism results in a plastic failure. (C) 2014 AIP Publishing LLC.

Design and Synthesis of New Benzimidazole-Carbazole Conjugates for the Stabilization of Human Telomeric DNA, Telomerase Inhibition, and Their Selective Action on Cancer Cells

Cell-permeable small molecules that enhance the stability of the G-quadruplex (G4) DNA structures are currently among the most intensively pursued ligands for inhibition of the telomerase activity. Herein we report the design and syntheses of four novel benzimidazole carbazole conjugates and demonstrate their high binding affinity to G4 DNA. Si nuclease assay confirmed the ligand mediated G-quadruplex DNA protection. Additional evidence from Telomeric Repeat Amplification Protocol (TRAP-LIG) assay demonstrated efficient telomerase inhibition activity by the ligands. Two of the ligands showed IC50 values in the sub-micromolar range in the TRAP-LIG assay, which are the best among the benzimidazole derivatives reported so far. The ligands also exhibited cancer cell selective nuclear internalization, nuclear condensation, fragmentation, and eventually antiproliferative activity in long-term cell viability assays. Annexin V-FITC/PI staining assays confirm that the cell death induced by the ligands follows an apoptotic pathway. An insight into the mode of ligand binding was obtained from the molecular dynamics simulations.

Laterally structured ripple and square phases with one and two dimensional thickness modulations in a model bilayer system

Molecular dynamics simulations of bilayers in a surfactant/co-surfactant/water system with explicit solvent molecules show formation of topologically distinct gel phases depending upon the bilayer composition. At low temperatures, the bilayers transform from the tilted gel phase, L beta', to the one dimensional (1D) rippled, P beta' phase as the surfactant concentration is increased. More interestingly, we observe a two dimensional (2D) square phase at higher surfactant concentration which, upon heating, transforms to the gel L beta' phase. The thickness modulations in the 1D rippled and square phases are asymmetric in two surfactant leaflets and the bilayer thickness varies by a factor of similar to 2 between maximum and minimum. The 1D ripple consists of a thinner interdigitated region of smaller extent alternating with a thicker non-interdigitated region. The 2D ripple phase is made up of two superimposed square lattices of maximum and minimum thicknesses with molecules of high tilt forming a square lattice translated from the lattice formed with the thickness minima. Using Voronoi diagrams we analyze the intricate interplay between the area-per-head-group, height modulations and chain tilt for the different ripple symmetries. Our simulations indicate that composition plays an important role in controlling the formation of low temperature gel phase symmetries and rippling accommodates the increased area-per-head-group of the surfactant molecules.

Local Structural and Environmental Factors Define the Efficiency of an RNA Pseudoknot Involved in Programmed Ribosomal Frameshift Process

In programmed -1 ribosomal frameshift, an RNA pseudoknot stalls the ribosome at specific sequence and restarts translation in a new reading frame. A precise understanding of structural characteristics of these pseudoknots and their PRF inducing ability has not been clear to date. To investigate this phenomenon, we have studied various structural aspects of a -1 PRF inducing RNA pseudoknot from BWYV using extensive molecular dynamics simulations. A set of functional and poorly functional forms, for which previous mutational data were available, were chosen for analysis. These structures differ from each other by either single base substitutions or base-pair replacements from the native structure. We have rationalized how certain mutations in RNA pseudoknot affect its function; e.g., a specific base substitution in loop 2 stabilizes the junction geometry by forming multiple noncanonical hydrogen bonds, leading to a highly rigid structure that could effectively resist ribosome-induced unfolding, thereby increasing efficiency. While, a CG to AU pair substitution in stem 1 leads to loss of noncanonical hydrogen bonds between stems and loop, resulting in a less stable structure and reduced PRF inducing ability, inversion of a pair in stem 2 alters specific base-pair geometry that might be required in ribosomal recognition of nucleobase groups, negatively affecting pseudoknot functioning. These observations illustrate that the ability of an RNA pseudoknot to induce -1 PRF with an optimal rate depends on several independent factors that contribute to either the local conformational variability or geometry

Ligand 5,10,15,20-Tetra(N-methyl-4-pyridyl)porphine (TMPyP4) Prefers the Parallel Propeller-Type Human Telomeric G-Quadruplex DNA over Its Other Polymorphs

The binding of ligand 5,10,15,20-tetra(N-methyl-4-pyridyl)porphine (TMPyP4) with telomeric and genomic G-quadruplex DNA has been extensively studied. However, a comparative study of interactions of TMPyP4 with different conformations of human telomeric G-quadruplex DNA, namely, parallel propeller-type (PP), antiparallel basket-type (AB), and mixed hybrid-type (MH) G-quadruplex DNA, has not been done. We considered all the possible binding sites in each of the G-quadruplex DNA structures and docked TMPyP4 to each one of them. The resultant most potent sites for binding were analyzed from the mean binding free energy of the complexes. Molecular dynamics simulations were then carried out, and analysis of the binding free energy of the TMPyP4-G-quadruplex complex showed that the binding of TMPyP4 with parallel propeller-type G-quadruplex DNA is preferred over the other two G-quadruplex DNA conformations. The results obtained from the change in solvent excluded surface area (SESA) and solvent accessible surface area (SASA) also support the more pronounced binding of the ligand with the parallel propeller-type G-quadruplex DNA.

Driving force of water entry into hydrophobic channels of carbon nanotubes: entropy or energy?

Spontaneous entry of water molecules inside single-wall carbon nanotubes (SWCNTs) has been confirmed by both simulations and experiments. Using molecular dynamics simulations, we have studied the thermodynamics of filling of a (6,6) carbon nanotube in a temperature range from 273 to 353K and with different strengths of the nanotube-water interaction. From explicit energy and entropy calculations using the two-phase thermodynamics method, we have presented a thermodynamic understanding of the filling behaviour of a nanotube. We show that both the energy and the entropy of transfer decrease with increasing temperature. On the other hand, scaling down the attractive part of the carbon-oxygen interaction results in increased energy of transfer while the entropy of transfer increases slowly with decreasing the interaction strength. Our results indicate that both energy and entropy favour water entry into (6,6) SWCNTs. Our results are compared with those of several recent studies of water entry into carbon nanotubes.

Spatial modulation of the composition of a binary liquid near a repulsive wall

When a binary liquid is confined by a strongly repulsive wall, the local density is depleted near the wall and an interface similar to that between the liquid and its vapor is formed. This analogy suggests that the composition of the binary liquid near this interface should exhibit spatial modulation similar to that near a liquid-vapor interface even if the interactions of the wall with the two components of the liquid are the same. The Guggenheim adsorption relation quantifies the concentrations of two components of a binary mixture near a liquid-vapor interface and qualitatively states that the majority (minority) component enriches the interface for negative (positive) mixing energy if the surface tensions of the two components are not very different. From molecular dynamics simulations of binary mixtures with different compositions and interactions we find that the Guggenheim relation is qualitatively satisfied at wall-induced interfaces for systems with negative mixing energy at all state points considered. For systems with positive mixing energy, this relation is found to be qualitatively valid at low densities, while it is violated at state points with high density where correlations in the liquid are strong. This observation is validated by a calculation of the density profiles of the two components of the mixture using density functional theory with the Ramakrishnan-Yussouff free-energy functional. Possible reasons for the violation of the Guggenheim relation are discussed.

Crack propagation in graphene

The crack initiation and growth mechanisms in an 2D graphene lattice structure are studied based on molecular dynamics simulations. Crack growth in an initial edge crack model in the arm-chair and the zig-zag lattice configurations of graphene are considered. Influence of the time steps on the post yielding behaviour of graphene is studied. Based on the results, a time step of 0.1 fs is recommended for consistent and accurate simulation of crack propagation. Effect of temperature on the crack propagation in graphene is also studied, considering adiabatic and isothermal conditions. Total energy and stress fields are analyzed. A systematic study of the bond stretching and bond reorientation phenomena is performed, which shows that the crack propagates after significant bond elongation and rotation in graphene. Variation of the crack speed with the change in crack length is estimated. (C) 2015 AIP Publishing LLC.

Suspensions of polymer-grafted nanoparticles with added polymers-Structure and effective pair-interactions

We present the results of combined experimental and theoretical (molecular dynamics simulations and integral equation theory) studies of the structure and effective interactions of suspensions of polymer grafted nanoparticles (PGNPs) in the presence of linear polymers. Due to the absence of systematic experimental and theoretical studies of PGNPs, it is widely believed that the structure and effective interactions in such binary mixtures would be very similar to those of an analogous soft colloidal material-star polymers. In our study, polystyrene-grafted gold nanoparticles with functionality f = 70 were mixed with linear polystyrene (PS) of two different molecular weights for obtaining two PGNP: PS size ratios, xi = 0.14 and 2.76 (where, xi = M-g/M-m, M-g and M-m being the molecular weights of grafting and matrix polymers, respectively). The experimental structure factor of PGNPs could be modeled with an effective potential (Model-X), which has been found to be widely applicable for star polymers. Similarly, the structure factor of the blends with xi = 0.14 could be modeled reasonably well, while the structure of blends with xi = 2.76 could not be captured, especially for high density of added polymers. A model (Model-Y) for effective interactions between PGNPs in a melt of matrix polymers also failed to provide good agreement with the experimental data for samples with xi = 2.76 and high density of added polymers. We tentatively attribute this anomaly in modeling the structure factor of blends with xi = 2.76 to the questionable assumption of Model-X in describing the added polymers as star polymers with functionality 2, which gets manifested in both polymer-polymer and polymer-PGNP interactions especially at higher fractions of added polymers. The failure of Model-Y may be due to the neglect of possible many-body interactions among PGNPs mediated by matrix polymers when the fraction of added polymers is high. These observations point to the need for a new framework to understand not only the structural behavior of PGNPs but also possibly their dynamics and thermo-mechanical properties as well. (C) 2015 AIP Publishing LLC.

Short-Time Beta Relaxation in Glass-Forming Liquids Is Cooperative in Nature

Temporal relaxation of density fluctuations in supercooled liquids near the glass transition occurs in multiple steps. Using molecular dynamics simulations for three model glass-forming liquids, we show that the short-time beta relaxation is cooperative in nature. Using finite-size scaling analysis, we extract a growing length scale associated with beta relaxation from the observed dependence of the beta relaxation time on the system size. We find, in qualitative agreement with the prediction of the inhomogeneous mode coupling theory, that the temperature dependence of this length scale is the same as that of the length scale that describes the spatial heterogeneity of local dynamics in the long-time alpha-relaxation regime.

Folding of Protein L with Implications for Collapse in the Denatured State Ensemble

A fundamental question in protein folding is whether the coil to globule collapse transition occurs during the initial stages of folding (burst phase) or simultaneously with the protein folding transition. Single molecule fluorescence resonance energy transfer (FRET) and small-angle X-ray scattering (SAXS) experiments disagree on whether Protein L collapse transition occurs during the burst phase of folding. We study Protein L folding using a coarse-grained model and molecular dynamics simulations. The collapse transition in Protein L is found to be concomitant with the folding transition. In the burst phase of folding, we find that FRET experiments overestimate radius of gyration, R-g, of the protein due to the application of Gaussian polymer chain end-to-end distribution to extract R-g from the FRET efficiency. FRET experiments estimate approximate to 6 angstrom decrease in R-g when the actual decrease is approximate to 3 angstrom on guanidinium chloride denaturant dilution from 7.5 to 1 M, thereby suggesting pronounced compaction in the protein dimensions in the burst phase. The approximate to 3 angstrom decrease is close to the statistical uncertainties of the R-g data measured from SAXS experiments, which suggest no compaction, leading to a disagreement with the FRET experiments. The transition-state ensemble (TSE) structures in Protein L folding are globular and extensive in agreement with the Psi-analysis experiments. The results support the hypothesis that the TSE of single domain proteins depends on protein topology and is not stabilized by local interactions alone.

Atomistic Simulation of Stacked Nucleosome Core Particles: Tail Bridging, the H4 Tail, and Effect of Hydrophobic Forces

We report the first atomistic simulation of two stacked nucleosome core particles (NCPs), with an aim to understand, in molecular detail, how they interact, the effect of salt concentration, and how different histone tails contribute to their interaction, with a special emphasis on the H4 tail, known to have the largest stabilizing effect on the NCP-NCP interaction. We do not observe specific K16-mediated interaction between the H4 tail and the H2A-H2B acidic patch, in contrast with the findings from crystallographic studies, but find that the stacking was stable even in the absence of this interaction. We perform simulations with the H4 tail (partially/completely) removed and find that the region between LYS-16 and LYS-20 of the H4 tail holds special importance in mediating the inter-NCP interaction. Performing similar tail-clipped simulations with the H3 tail removed, we compare the roles of the H3 and H4 tails in maintaining the stacking. We discuss the relevance of our simulation results to the bilayer and other liquid-crystalline phases exhibited by NCPs in vitro and, through an analysis of the histone-histone interface, identify the interactions that could possibly stabilize the inter-NCP interaction in these columnar mesophases. Through the mechanical disruption of the stacked nucleosome system using steered molecular dynamics, we quantify the strength of inter-NCP stacking in the presence and absence of salt. We disrupt the stacking at some specific sites of internucleosomal tail-DNA contact and perform a comparative quantification of the binding strengths of various tails in stabilizing the stacking. We also examine how hydrophobic interactions may contribute to the overall stability of the stacking and find a marked difference in the role of hydrophobic forces as compared with electrostatic forces in determining the stability of the stacked nucleosome system.

Self-Organization of n-Alkane Chains in Water: Length Dependent Crossover from Helix and Toroid to Molten Globule

We demonstrate a chain length dependent crossover in the structural properties of linear hydrocarbon (n-alkane) chains using detailed atomistic simulations in explicit water. We identify a number of exotic structures of the polymer chain through energy minimization of representative snapshots collected from molecular dynamics trajectory. While the collapsed state is ring-like (circular) for small chains (CnH2n+2; n <= 20) and spherical for very long ones (n = 100), we find the emergence of ordered helical structures at intermediate lengths (n similar to 40). We find different types of disordered helices and toroid-like structures at n = 60. We also report a sharp transition in the stability of the collapsed state as a function of the chain length through relevant free energy calculations. While the collapsed state is only marginally metastable for C20H42, a clear bistable free energy surface emerges only when the chain is about 30 monomers long. For n = 30, the polymer exhibits an intermittent oscillation between the collapsed and the coil structures, characteristic of two stable states separated by a small barrier.