Ligand dependent intra and inter subunit communication in human tryptophanyl tRNA synthetase as deduced from the dynamics of structure networks

Homodimeric protein tryptophanyl tRNA synthetase (TrpRS) has a Rossmann fold domain and belongs to the 1c subclass of aminoacyl tRNA synthetases. This enzyme performs the function of acylating the cognate tRNA. This process involves a number of molecules (2 protein subunits, 2 tRNAs and 2 activated Trps) and thus it is difficult to follow the complex steps in this process. Structures of human TrpRS complexed with certain ligands are available. Based on structural and biochemical data, mechanism of activation of Trp has been speculated. However, no structure has yet been solved in the presence of both the tRNA(Trp) and the activated Trp (TrpAMP). In this study, we have modeled the structure of human TrpRS bound to the activated ligand and the cognate tRNA. In addition, we have performed molecular dynamics (MD) simulations on these models as well as other complexes to capture the dynamical process of ligand induced conformational changes. We have analyzed both the local and global changes in the protein conformation from the protein structure network (PSN) of MD snapshots, by a method which was recently developed in our laboratory in the context of the functionally monomeric protein, methionyl tRNA synthetase. From these investigations, we obtain important information such as the ligand induced correlation between different residues of this protein, asymmetric binding of the ligands to the two subunits of the protein as seen in the crystal structure analysis, and the path of communication between the anticodon region and the aminoacylation site. Here we are able to elucidate the role of dimer interface at a level of detail, which has not been captured so far.

Vitamin A and thyroxine carrier proteins in chicken plasma Steady-state control of the plasma level of free retinol-binding protein and free thyroxine

1. 1. The binding parameters of prealbumin-2 with retinol-binding protein and thyroxine (T4) revealed the existence of distinct and multiple sites for both retinol-binding protein and T4. 2. 2. From the analysis of binding parameters of retinol-binding protein with prealbumin-2 it is clear that under steady-state conditions about 99% of the holo-retinol-binding protein remains bound to prealbumin-2. 3. 3. Equilibrium dialysis studies on binding properties of thyroid hormones with prealbumin-2 revealed that it has a single high affinity site and three low affinity sites. 4. 4. The occurrence of three carrier proteins for thyroid hormones, thyroxine-binding globulin, prealbumin-2 and albumin has been demonstrated. However, the chicken thyroxine-binding globulin differs from human thyroxine-binding globulin by being relatively less acidic and occuring at a two-fold lower concentration. But the thyroid hormone binding parameters are comparable. 5. 5. Highly sensitive methods were developed for determination of T4 binding capacities of the various proteins and plasma level of total T4 by fractionation of carrier proteins and further quantitatively employing in electrophoresis and equilibrium dialysis. 6. 6. The thyroxine-binding proteins were found to be two types, one (viz., thyroxine-binding globulin) of great affinity but of low binding capacity, which mainly acts as reservoir of T4, and another (viz.,prealbumin-2) of low affinity but of high binding capacity, which can participate predominantly in the control of the free T4 pool.

The level of sequestered calcium in vegetative amoebae of Dictyostelium discoideum can predict post-aggregative cell fate

When freshly starved amoebae of Dictyostelium discoideum are stained with chlortetracycline (CTC), a cell type-specific fluorescent probe for membrane-associated calcium (Ca2+) the resulting fluorescence distribution falls into two functional classes. Fluorescence-activated cell sorting shows that highly fluorescing amoebae tend to enter the prestalk pathway while those with low fluorescence tend to become prespores. In the light of previous findings, these results indicate that in addition to cell cycle phase at starvation, phenotypic variation in the level of sequestered calcium is an early correlate of cell fate.

Influence of the highest occupied molecular orbital energy level of the donor material on the effectiveness of the anode buffer layer in organic solar cells

Efficiency of organic photovoltaic cells based on organic electron donor/organic electron acceptor junctions can be strongly improved when the transparent conductive Anode is coated with a Buffer Layer (ABL). Here, the effects of a metal (gold) or oxide (molybdenum oxide) ABL are reported, as a function of the Highest Occupied Molecular Orbital (HOMO) of different electron donors. The results indicate that a good matching between the work function of the anode and the highest occupied molecular orbital of the donor material is the major factor limiting the hole transfer efficiency. Indeed, gold is efficient as ABL only when the HOMO of the organic donor is close to its work function Phi(Au). Therefore we show that the MoO(3) oxide has a wider field of application as ABL than gold. (C) 2011 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim

Influence of planning and governance on the level of urban services

A theoretical framework to analyse the interaction of planning and governance on the extent of outgrowth and level of services is proposed. An indicator framework for quantifying sprawl is also proposed and the same is operationalised for Bangalore. The indicators comprise spatial metrics (derived from temporal satellite remote sensing data) and other metrics obtained from a house-hold survey. The interaction of different indicators with respect to the core city and the outgrowth is determined by multi-dimensional scaling. The analysis reveals the underlying similarities (and dissimilarities) that relate with the different governance structures that prevail here. The paper concludes outlining the challenges in addressing urban sprawl while ensuring adequate level of services that planning and governance have to ensure towards achieving sustainable urbanisation.

Qualitative toxicity assessment of silver nanoparticles on the fresh water bacterial isolates and consortium at low level of exposure concentration

Silver nanoparticles (AgNPs) pose a high risk of exposure to the natural environment owing to their extensive usage in various consumer products. In the present study we attempted to understand the harmful effect of AgNPs at environmentally relevant low concentration levels (<= 1 ppm) towards two different freshwater bacterial isolates and their consortium. The standard plate count assay suggested that the AgNPs were toxic towards the fresh water bacterial isolates as well as the consortium, though toxicity was significantly reduced for the cells in the consortium. The oxidative stress assessment and membrane permeability studies corroborated with the toxicity data. The detailed electron microscopic studies suggested the cell degrading potential of the AgNPs, and the FT-IR studies confirmed the involvement of the surface groups in the toxic effects. No significant ion leaching from the AgNPs was observed at the applied concentration levels signifying the dominant role of the particle size, and size distribution in bacterial toxicity. The reduced toxicity for the cells in the consortium than the individual isolates has major significance in further studies on the ecotoxicity of the AgNPs. (C) 2014 Elsevier Inc. All rights reserved.

BIOCARDS AND LEVEL OF ABSTRACTION

Biocards are formal descriptions of biological phenomena and their underlying functional principles. They are used in bioinspired design to document search results and to communicate the findings for use in the further design process. The present study explored the effect of abstraction level used in biocards. This was done in two workshops conducted with design students in Denmark and India. Students were given a design assignment and instructions for how to perform the BID ideation work. Half of the students were given biocards with abstract descriptions while the other half got biocards with concrete descriptions. The novelty of found solutions was evaluated by the students by rating novelty of each solution on a scale from 1 to 5. Mean values for abstract descriptions were 0,3 higher than for concrete descriptions indicating that more innovative solutions were found when students used biocards with abstract descriptions compared to concrete descriptions. The difference in mean value is significant with a confidence level better than 1%. It seems likely that more abstract descriptions in biocards helps avoiding design fixation in biomimetic design work.

Molecular dynamics and simulations study on the vibrational and electronic solvatochromism of benzophenone

Solvent plays a key role in diverse physico-chemical and biological processes. Therefore, understanding solute-solvent interactions at the molecular level of detail is of utmost importance. A comprehensive solvatochromic analysis of benzophenone (Bzp) was carried out in various solvents using Raman and electronic spectroscopy, in conjunction with Density Functional Theory (DFT) calculations of supramolecular solute-solvent clusters generated using classical Molecular Dynamics Simulations (c-MDSs). The >C=O stretching frequency undergoes a bathochromic shift with solvent polarity. Interestingly, in protic solvents this peak appears as a doublet: c-MDS and ad hoc explicit solvent ab initio calculations suggest that the lower and higher frequency peaks are associated with the hydrogen bonded and dangling carbonyl group of Bzp, respectively. Additionally, the dangling carbonyl in methanol (MeOH) solvent is 4 cm(-1) blue-shifted relative to acetonitrile solvent, despite their similar dipolarity/polarizability. This suggests that the cybotactic region of the dangling carbonyl group in MeOH is very different from its bulk solvent structure. Therefore, we propose that this blue-shift of the dangling carbonyl originates in the hydrophobic solvation shell around it resulting from extended hydrogen bonding network of the protic solvents. Furthermore, the 1(1)n pi* (band I) and 1(1)pi pi* (band II) electronic transitions show a hypsochromic and bathochromic shift, respectively. In particular, these shifts in protic solvents are due to differences in their excited state-hydrogen bonding mechanisms. Additionally, a linear relationship is obtained for band I and the >C=O stretching frequency (cm(-1)), which suggests that the different excitation wavelengths in band I correspond to different solvation states. Therefore, we hypothesize that the variation in excitation wavelengths in band I could arise from different solvation states leading to varying solvation dynamics. This will have implications for ultrafast processes associated with electron-transfer, charge transfer, and also the photophysical aspects of excited states. (C) 2016 AIP Publishing LLC.

Stability of large flexible damped spacecraft modeled as elastic continua

Vibrational stability of a large flexible, structurally damped spacecraft subject to large rigid body rotations is analysed modelling the system as an elastic continuum. Using solution of rigid body attitude motion under torque free conditions and modal analysis, the vibrational equations are reduced to ordinary differential equations with time-varying coefficients. Stability analysis is carried out using Floquet theory and Sonin-Polya theorem. The cases of spinning and non-spinning spacecraft idealized as a flexible beam plate undergoing simple structural vibration are analysed in detail. The critical damping required for stabilization is shown to be a function of the spacecraft's inertia ratio and the level of disturbance.

Five-level inverter scheme for an open-end winding induction machine with less number of switches

Higher level of inversion is achieved with a less number of switches in the proposed scheme. The scheme proposes a five-level inverter for an open-end winding induction motor which uses only two DC-link rectifiers of voltage rating of Vdc/4, a neutral-point clamped (NPC) three-level inverter and a two-level inverter. Even though the two-level inverter is connected to the high-voltage side, it is always in square-wave operation. Since the two-level inverter is not switching in a pulse width modulated fashion and the magnitude of switching transient is only half compared to the convention three-level NPC inverter, the switching losses and electromagnetic interference is not so high. The scheme is experimentally verified on a 2.5 kW induction machine.

Baculovirus mediated high-level expression of luciferase in silkworm cells and larvae

Bombyx mori nuclear polyhedrosis virus (BmNPV)-based baculovirus expression system exploits silkworm larvae as an economical alternative to large-scale cell cultures for production of biomolecules. To generate recombinant BmNPV at high efficiency, we have achieved high efficiency transfection of B. mori cells, BmN, through lipofection. Optimal conditions for lipofection were standardized by quantification of the transient expression level of firefly luciferase (luc) reporter gene under control of an immediate early gene promoter of BmNPV Lipofection was 50-fold and 100-fold more efficient than the calcium phosphate method for transfecting BmN and Sf9 cells, respectively. Lipofection enabled us to generate a recombinant BmNPV (vBmluc), harboring luc under control of the strong polyhedrin promoter On infection with vBmluc, luciferase was expressed at very high levels, 170 mu g/10(6) BmN cells or 13 mg/larva. Expression of luciferase in vBmluc-infected larvae was visualized by luminescence emission instantaneously following luciferin injection generating ''glowing silkworms''.

Reconstructing Solid Model from 2D Scanned Images of Biological Organs for Finite Element Simulation

This work presents a methodology to reconstruct 3D biological organs from image sequences or other scan data using readily available free softwares with the final goal of using the organs (3D solids) for finite element analysis. The methodology deals with issues such as segmentation, conversion to polygonal surface meshes, and finally conversion of these meshes to 3D solids. The user is able to control the detail or the level of complexity of the solid constructed. The methodology is illustrated using 3D reconstruction of a porcine liver as an example. Finally, the reconstructed liver is imported into the commercial software ANSYS, and together with a cyst inside the liver, a nonlinear analysis performed. The results confirm that the methodology can be used for obtaining 3D geometry of biological organs. The results also demonstrate that the geometry obtained by following this methodology can be used for the nonlinear finite element analysis of organs. The methodology (or the procedure) would be of use in surgery planning and surgery simulation since both of these extensively use finite elements for numerical simulations and it is better if these simulations are carried out on patient specific organ geometries. Instead of following the present methodology, it would cost a lot to buy a commercial software which can reconstruct 3D biological organs from scanned image sequences.

Metabolites of PPI-2458, a selective, irreversible inhibitor of methionine aminopeptidase-2: structure determination and In vivo activity

The natural product fumagillin exhibits potent antiproliferative and antiangiogenic properties. The semisynthetic analog PPI-2458, (3R,4S,5S,6R)-5-methoxy-4-(2R,3R)-2-methyl-3-(3-methylbut-2-enyl) oxiran-2-yl]-1-oxaspiro2.5]octan-6-yl] N-(2R)-1-amino-3-methyl-1-oxobutan-2-yl]carbamate, demonstrates rapid inactivation of its molecular target, methionine aminopeptidase-2 (MetAP2), and good efficacy in several rodent models of cancer and inflammation with oral dosing despite low apparent oral bioavailability. To probe the basis of its in vivo efficacy, the metabolism of PPI-2458 was studied in detail. Reaction phenotyping identified CYP3A4/5 as the major source of metabolism in humans. Six metabolites were isolated from liver microsomes and characterized by mass spectrometry and nuclear resonance spectroscopy, and their structures were confirmed by chemical synthesis. The synthetic metabolites showed correlated inhibition of MetAP2 enzymatic activity and vascular endothelial cell growth. In an ex vivo experiment, MetAP2 inhibition in white blood cells, thymus, and lymph nodes in rats after single dosing with PPI-2458 and the isolated metabolites was found to correlate with the in vitro activity of the individual species. In a phase 1 clinical study, PPI-2458 was administered to patients with non-Hodgkin lymphoma. At 15 mg administered orally every other day, MetAP2 in whole blood was 80% inactivated for up to 48 hours, although the exposure of the parent compound was only similar to 10% that of the summed cytochrome P450 metabolites. Taken together, the data confirm the participation of active metabolites in the in vivo efficacy of PPI-2458. The structures define a metabolic pathway for PPI-2458 that is distinct from that of TNP-470 ((3R, 4S, 5S, 6R)-5-methoxy-4-(2R, 3R)-2-methyl-3-(3-methylbut-2-enyl)oxiran-2-yl]-1-oxaspiro2.5]octan-6 -yl] N-(2-chloroacetyl)carbamate). The high level of MetAP2 inhibition achieved in vivo supports the value of fumagillin-derived therapeutics for angiogenic diseases.

A fluorescent molecularly-imprinted polymer gate with temperature and pH as inputs for detection of alpha-fetoprotein

In this work, we have reported a new approach on the use of stimuli-responsive molecularly imprinted polymer (MIP) for trace level sensing of alpha-fetoprotein (AFP), which is a well know cancer biomarker. The stimuli-responsive MIP is composed of three components, a thermo-responsive monomer, a pH responsive component (tyrosine derivative) and a highly fluorescent vinyl silane modified carbon dot. The synthesized AFP-imprinted polymer possesses excellent selectivity towards their template molecule and dual-stimuli responsive behavior. Along with this, the imprinted polymer was also explored as `OR' logic gate with two stimuli (pH and temperature) as inputs. However, the non-imprinted polymers did not have such `OR' gate property, which confirms the role of template binding. The imprinted polymer was also used for estimation of AFP in the concentration range of 3.96-80.0 ng mL(-1), with limit of detection (LOD) 0.42 ng mL(-1). The role of proposed sensor was successfully exploited for analysis of AFP in real human blood plasma, serum and urine sample. (C) 2015 Elsevier B.V. All rights reserved.