Morphology and Chemical Composition of Magnetic Particles Separated from Coal Fly Ash

Iron and other metal compounds are the materials that often appear in coal seams, because they also appear as a component of former organic matter in coal rocks. Although iron is the dominant element in coal rocks, other metals such as titanium, lead, cobalt, nickel, and copper are also present. In this study, the properties of magnetic particles of a size between 1 and 20 µm of globular structure and iron containing, were separated from coal fly ash, and studied using a scanning electron microscopy, energy disperse spectroscopy, and X-ray diffraction spectroscopy. The investigations were comprised of micrographs of the structure of these particles, their elemental composition, and phase analysis.

Cooling Curve Analysis of A356 Alloy by Conventional Casting and the Effect of Stirring

The present investigation attempted to explore the effect of stirring during solidification of Aluminum A356 alloy, mainly focusing on the change from dendrite to globular structure. For this purpose samples of A356 alloy were melted in the electrical resistance furnace and cooling curves were recorded for each level agitation. The experimental curves were numerically processed by calculating first and second derivatives. From these were determined temperatures and times of start nucleation of alpha solid and eutectic reaction.

Characterization of gliadin, secalin and hordein fractions using analytical techniques

Prolamins, alcohol soluble storage proteins of the Triticeae tribe of Gramineae family, are known as gliadin, secalin and hordein in wheat, rye and barley respectively. Prolamins were extracted from fifteen cultivars using DuPont protocol to study their physiochemical, morphological and structural characteristics. SDS-PAGE of prolamins showed well resolved low molecular weight proteins with significant amount of albumin and globulin as cross-contaminant. The β-sheet (32.72–37.41%) and β-turn (30.36–37.91%) were found higher in gliadins, while α-helix (20.32–28.95%) and random coil (9.05–10.28%) in hordeins. The high colloidal stability as depicted by zeta-potential was observed in gliadins (23.5–27.0 mV) followed secalins (11.2–16.6 mV) and hordeins (4.1–7.8 mV). Surface morphology by SEM illustrated the globular particle arrangement in gliadins, sheet like arrangement in secalins and stacked flaky particle arrangement in hordeins fraction. TEM studies showed that secalin and hordein fractions were globular in shape while gliadins in addition to globular structure also possessed rod-shaped particle arrangement. XRD pattern of prolamin fractions showed the ordered crystalline domain at 2θ values of 44.1°, 37.8° and 10.4°. The extracted prolamins fractions showed amorphous as well as crystalline structures as revealed by XRD and TEM analysis. Space saving hexagonal molecular symmetry was also observed in TEM molecular arrangement of prolamins which has profound application in development of plant-based polymers and fibres.

LRET-derived HADDOCK structural models describe the conformational heterogeneity required for processivity of the Mre11-Rad50 DNA damage repair complex

The Mre11-Rad50-Nbs1 protein complex is one of the first responders to DNA double strand breaks. Studies have shown that the catalytic activities of the evolutionarily conserved Mre11-Rad50 (MR) core complex depend on an ATP-dependent global conformational change that takes the macromolecule from an open, extended structure in the absence of ATP to a closed, globular structure when ATP is bound. We have previously identified an additional 'partially open' conformation using Luminescence Resonance Energy Transfer (LRET) experiments. Here, a combination of LRET and the molecular docking program HADDOCK was used to further investigate this partially open state and identify three conformations of ATP-bound MR in solution: closed, partially open, and open, which are in addition to the extended, apo conformation. These models are supported with mutagenesis and SAXS data that corroborate the presence of these three states and suggest a mechanism for the processivity of the MR complex along the DNA.

1H, 13C, 15N backbone resonance assignment for the 1–164 construct of human XRCC4

DNA double-strand breaks (DSBs) represent the most cytotoxic DNA lesions, as—if mis- or unrepaired—they can cause cell death or lead to genome instability, which in turn can cause cancer. DSBs are repaired by two major pathways termed homologous recombination and non-homologous end-joining (NHEJ). NHEJ is responsible for repairing the vast majority of DSBs arising in human cells. Defects in NHEJ factors are also associated with microcephaly, primordial dwarfism and immune deficiencies. One of the key proteins important for mediating NHEJ is XRCC4. XRCC4 is a dimer, with the dimer interface mediated by an extended coiled-coil. The N-terminal head domain forms a mixed alpha–beta globular structure. Numerous factors interact with the C-terminus of the coiled-coil domain, which is also associated with significant self-association between XRCC4 dimers. A range of construct lengths of human XRCC4 were expressed and purified, and the 1–164 variant had the best NMR properties, as judged by consistent linewidths, and chemical shift dispersion. In this work we report the 1H, 15 N and 13C backbone resonance assignments of human XRCC4 in the solution form of the 1–164 construct. Assignments were obtained by heteronuclear multidimensional NMR spectroscopy. In total, 156 of 161 assignable residues of XRCC4 were assigned to resonances in the TROSY spectrum, with an additional 11 resonances assigned to His-Tag residues. Prediction of solution secondary structure from a chemical shift analysis using the TALOS + webserver is in good agreement with the published X-ray crystal structures of this protein.

SANS STUDIES ON THE BOVINE SERUM ALBUMIN DENATURATION IN THE PRESENCE OF SDS

The effect of the presence of sodium dodecyl sulfate (SDS) on the denaturation of bovine serum albumin (BSA) has been studied using 36 m small-angle neutron scattering (SANS) BATAN spectrometer (SMARTer). The neutron scattering data reduction used the Graphical Reduction and Analysis SANS Program (GRASP) software, and the fitting process used the IGOR SANS Analysis software. The denaturation process was identified by observing the changes BSA globular structure. The experimental results showed the addition of SDS at low concentrations (2 mM, 5 mM, 10 mM) into BSA solution at pH 7 do not cause a significant change in the size of the BSA globular structure. The SANS scattering profile of BSA fitted with the triaxial ellipsoid model, a simple shape approach for protein globular structure. The fitting result showed the semi-axis B for BSA in the addition of 2 mM, 5 mM, 10 mM SDS were 33.8 Å, 33.8 Å, and 37.8 Å, respectively. While the semi-axis A and semi-axis C were constant for those three variations at 14.6 Å and 32.2 Å, respectively. In higher addition of SDS, the globular structure of BSA unfolded into flexible cylinder structure with the radius of 14.4 Å and length of 83.5 Å. The denaturation of BSA was clearly showed by the addition of 40 mM SDS. The structure of BSA in this condition fitted to fractal structure with fractal dimension of 1.1, the block radius of 16.7 Å and the correlation length of 42.5 Å. These results indicated that the addition of SDS at low concentrations has not caused the denaturation of BSA. Meanwhile, the addition of SDS at high concentrations made BSA to unfold that lead to the denaturation of BSA.

Chemical characterization (LC–MS–ESI), cytotoxic activity and intracellular localization of PAMAM G4 in leukemia cells

Generation 4 of polyamidoamine dendrimer (G4-PAMAM) has several biological effects due to its tridimensional globular structure, repetitive branched amides, tertiary amines, and amino-terminal subunit groups liked to a common core. G4-PAMAM is cytotoxic due to its positive charges. However, its cytotoxicity could increase in cancer cells due to the excessive intracellular negative charges in these cells. Furthermore, this work reports G4-PAMAM chemical structural characterization using UHPLC-QTOF-MS/MS (LC–MS) by electrospray ionization to measure its population according to its positive charges. Additionally, the antiproliferative effects and intracellular localization were explored in the HMC-1 and K-562 cell lines by confocal microscopy. The LC–MS results show that G4-PAMAM generated multivalent mass spectrum values, and its protonated terminal amino groups produced numerous positive charges, which allowed us to determine its exact mass despite having a high molecular weight. Additionally, G4-PAMAM showed antiproliferative activity in the HMC-1 tumor cell line after 24 h (IC50 = 16.97 µM), 48 h (IC50 = 7.02 µM) and 72 h (IC50 = 5.98 µM) and in the K-562 cell line after 24 h (IC50 = 15.14 µM), 48 h (IC50 = 14.18 µM) and 72 h (IC50 = 9.91 µM). Finally, our results showed that the G4-PAMAM dendrimers were located in the cytoplasm and nucleus in both tumor cell lines studied.

Divergence Entropy-Based Evaluation of Hydrophobic Core in Aggressive and Resistant Forms of Transthyretin

The two forms of transthyretin differing slightly in the tertiary structure, despite the presence of five mutations, show radically different properties in terms of susceptibility to the amyloid transformation process. These two forms of transthyretin are the object of analysis. The search for the sources of these differences was carried out by means of a comparative analysis of the structure of these molecules in their native and early intermediate stage forms in the folding process. The criterion for assessing the degree of similarity and differences is the status of the hydrophobic core. The comparison of the level of arrangement of the hydrophobic core and its initial stages is possible thanks to the application of divergence entropy for the early intermediate stage and for the final forms. It was shown that the minimal differences observed in the structure of the hydrophobic core of the forms available in PDB, turned out to be significantly different in the early stage (ES) structure in folding process. The determined values of divergence entropy for both ES forms indicate the presence of the seed of hydrophobic core only in the form resistant to amyloid transformation. In the form of aggressively undergoing amyloid transformation, the structure lacking such a seed is revealed, being a stretched one with a high content of β-type structure. In the discussed case, the active presence of water in the structural transformation of proteins expressed in the fuzzy oil drop model (FOD) is of decisive importance for the generation of the final protein structure. It has been shown that the resistant form tends to generate a centric hydrophobic core with the possibility of creating a globular structure, i.e. a spherical micelle-like form. The aggressively transforming form reveals in the structure of its early intermediate, a tendency to form the ribbon-like micelle as observed in amyloid.

Title: Characterization of gliadin, secalin and hordein using advance analytical techniques

Prolamins extracted from wheat, rye and barley cultivars were analysed for colour characteristics, SDS-PAGE, Amino Acid, Dynamic Light Scattering (DLS), Zeta Potential, Scanning Electron Microscopy (SEM), Energy Dispersive X-Ray (EDX), Transmission Electron Microscopy (TEM) and X-ray Diffraction (XRD) to elucidate the structure. Amino acid analysis showed significant variation among the prolamins and the predominant essential amino acids were found to be leucine, phenylalanine and valine whereas predominant non-essential amino acids were glutamic acid, arginine and aspartic acid. All the prolamins exhibited positive zeta-potential, however gliadin had a significantly higher zeta potential value. TEM studies of prolamins revealed the compact globular structure but gliadin also had rod-shaped structure. Morphology by SEM illustrated the globular particle arrangement in gliadin and sheet like arrangement in secalin and hordein. XRD pattern of prolamin showed the ordered crystalline domain of prolamin at 44.1°, 37.8° and 10.4°. The d-spacing obtained from XRD and TEM analysis also supports the crystalline domain of prolamin apart from amorphous domain.

A “spindle and thread”-mechanism unblocks translation of N-terminally disordered proteins

Protein disorder is a major hurdle for structural biology. A prominent example is the tumour suppressor p53, whose low expression levels and poor conformational stability due to a high degree of disorder pose major challenges to the development of cancer therapeutics. Here, we address these issues by fusing p53 to an engineered spider silk domain termed NT*. The chimeric protein displays highly efficient translation in vitro and in E. coli and is fully active in human cancer cells. The transmission electron microscopy structure and native mass spectrometry reveal that the full-length p53 fusion protein adopts a compact conformation. Molecular dynamics simulations show that the disordered transactivation domain of p53 is wound around the NT* domain via a series of folding events, resulting in a globular structure. We find that expression of B-Raf, another partially disordered cancer target, is similarly enhanced by fusion to NT*. In summary, we demonstrate how inducing co-translational folding via a molecular “spindle and thread” mechanism can overcome poor translation efficiency of partially disordered proteins.