Calorimetric study of mutant human lysozymes with partially introduced Ca2+ binding sites and its efficient refolding system from inclusion bodies

During the process of evolution, ancestral lysozymes evolved intocalcium-binding lysozymes by acquiring three critical aspartate residues atpositions 86, 91 and 92. To investigate the process of the acquisition ofcalcium-binding ability, two of the aspartates were partially introducedinto human lysozyme at positions 86, 91 and 92. These mutants (HLQ86D,HLA92D and HLQ86D/D91Q/A92D), having two critical aspartates incalcium-binding sites, were expressed in Escherichia coli as non-activeinclusion bodies. For the preparation of lysozyme samples, a refoldingsystem using thioredoxin was established. This system allowed for effectiverefolding of wild-type and mutant lysozymes, and 100% of activity wasrecovered within 4 days. The calcium ion dependence of the meltingtemperature (Tm) of wild-type and mutant lysozymes was investigated bydifferential scanning calorimetry at pH 4.5. The Tm values of wild-type,HLQ86D and HLA92D mutants were not dependent on calcium ion concentration.However, the Tm of HLQ86D/D91Q/A92D was 4 degrees higher in the presence of50 mM CaCl2 than in its absence, and the calcium-binding constant of thismutant was estimated to be 2.25(+/-0.25)x10(2) M(-1) at pH 4.5. Moreover,the calcium-binding ability of this mutant was confirmed by the resultusing Sephadex G-25 gel chromatography. These results indicate that it isindispensable to have at least two aspartates at positions 86 and 92 foracquisition of calcium-binding ability. The process of the acquisition ofcalcium-binding site during evolution of calcium-binding lysozyme isdiscussed.     

Effects of main particle diameter on improving particle flowability for compressed packing fraction in a smaller particle admixing system

Particle flowability can be improved by admixing particles smaller than the original particles (main particles). However, the mechanisms by which this technique improves flowability are not yet fully understood. In this study, we examined compressed packing in a particle bed, which is affected by particle flowability. To estimate the mechanism of improvement, we investigated the effects of the main particle diameter on the improvement of compressed packing fractions experimentally.The main particles were 397 and 1460 nm in diameter and the admixed particles were 8, 21, 62, and 104 nm in diameter. The main and admixed particles were mixed in various mass ratios, and the compressed packing fractions of the mixtures were measured. SEM images were used to analyze the coverage diameter and the surface coverage ratio of the admixed particles on the main particles. The main particle packing fraction was improved as the diameter ratio (=main particles/admixed particles) increased. This was explained by a linked rigid-3-bodies model with leverage. Furthermore, the actual surface coverage ratio at which the most improved packing fraction was obtained decreased with increasing main particle diameter. This was explained by the difference in the curvature of the main particle surface.     

Serum matrix metalloproteinase-3 in hemodialysis patients with dialysis-related amyloidosis

Background: Matrix metalloproteinase‐3 (MMP‐3) has been linked to osteoarticular destruction in rheumatic arthritis. To investigate the role of MMP‐3 in dialysis‐related amyloidosis (DRA), we determined serum MMP‐3 in long‐term hemodialysis (HD) patients with and without clinical manifestations of DRA. Methods: Thirty‐three subjects (63% female, 3% diabetic) enrolled in the study between September 2001 and June 2003. All patients underwent standard HD three times per week, using high‐flux dialyzers. Four patients had active DRA complications (DRA patients), whereas the others (n = 29) had no evidence of DRA. We determined serum concentrations of MMP‐3, C‐reactive protein (CRP), β₂‐microglobulin (β₂M), and interleukin‐6 (IL‐6). We also studied the effects of hemodiafiltration (HDF) on inflammatory measures by transferring the DRA patients from regular HD to predilution HDF. Results: The DRA group had been on dialysis significantly longer than the control group. Significant positive correlations were observed between MMP‐3 and IL‐6 (R² = 0.5143, p < 0.0001) and MMP‐3 and CRP (R² = 0.6492, p < 0.0001). IL‐6 levels increased after a single dialysis treatment, but this effect was minimal with predilution HDF (the increment of IL‐6 levels did not exceed 10 pg/mL). Serum MMP‐3 levels decreased in parallel with the decrease of IL‐6. Conclusions: MMP‐3 serum levels increase in accordance with clinical manifestations of DRA and elevated circulating levels of IL‐6. For the evaluation of the pathophysiologic state of DRA, serum MMP‐3 may be a useful predictor indicative of chronic inflammation and osteoarticular disorders in DRA patients.     

Morphological evolution of NiO-ScSZ composite in a high-temperature reducing atmosphere

The microstructural change in composite of nickel oxide and scandia-stabilized zirconia (NiO-ScSZ) under a reducing atmosphere was observed by scanning electron microscopy (SEM). The morphological transformation was noticeable after high-temperature treatment with the formation of two peculiar microstructures; i.e., fibrous zirconia and metallic nickel with wrinkled surface. It was suggested that partial reduction of the nickel species dissolved in ScSZ lattice triggered the formation of these characteristic morphologies. The growth of fibrous zirconia appeared to be promoted via interfacial reaction between the metallic Ni particles and the zirconia phase. The agglomeration of metallic nickel proceeded by the reduction at high temperatures and then the surface was transformed to the wrinkled morphology. The unique wrinkled pattern was often observed for the large agglomerated nickel particles. This drastic change in Ni-ScSZ microstructure upon the high-temperature reduction degraded the electrochemical performance of solid oxide fuel cells.     

Modification of transport properties of ion exchange membranes,XIII. Surface-modified cation exchange membranes prepared from polyamines and a copolymer membrane containing p-stryenesulfonyl chloride

A membrane precursor was prepared by the copolymerization of p-styrenesulfonyl chloride and divinylbenzene in the presence of a poly(vinyl chloride) supported cloth. The surface of the membrane was treated with various amines, and then the sulfonyl chloride groups in the internal part of the membrane were hydrolyzed with an aqueous sodium hydroxide solution in organic methanol or acetone solvent. Hydrolysis conditions were examined for the introduction of the sulfonic acid groups. The membrane modified by ethylenediamine exhibited slightly bipolar properties during the electrodialysis of seawater. Surface-modified membranes using tetraethylenepentamine exhibited excellent monovalent cation permselectivity, low membrane electric resistance, and long performance durability during the electrodialysis of seawater.     

Molecular Docking Reveals the Binding Modes of Anticancer Alkylphospholipids and Lysophosphatidylcholine within the Catalytic Domain of Cytidine Triphosphate: Phosphocholine Cytidyltransferase

Alkylphospholipids are synthetic analogues of endogenous phosphatidylcholines with a remarkable ability: induce the selective apoptosis of exponentially growing tumor cells. One hypothesis concerning their mechanism of action is the inhibition of cytidine triphosphate:phosphocholine cytidyltransferase (CCT), which would significantly suppress the phosphatidylcholine biosynthesis to trigger apoptosis. Herein, homology modeling, docking simulations, and the analyses of molecular interaction fields are used to suggest the most probable binding modes of four alkylphospholipids (edelfosine, erucylphosphocholine, perifosine, and miltefosine) and lysophosphatidylcholine at the catalytic domain of human CCT. All compounds display bind modes in agreement with the corresponding groups found in the CCT substrate, phosphocholine, while their binding strengths are increased because of the interaction of the alkyl chains with hydrophobic residues from the M domain of the protein. Analyses of the geometry of the CCT binding‐site also suggest that small groups, such as benzyl/2‐phenylethyl ethers or equivalent heterocycles, could replace the O‐methyl group in edelfosine to yield even better inhibitors. It is believed this study can guide the development of new alkylphospholipids with an improved profile for the inhibition of phosphatidylcholine biosynthesis, a critical component for cell cycle progression that can be explored in cancer chemotherapy. Practical Applications: Studies focusing on the interactions between small ligands and their protein targets are decisive for the comprehension of how conformational changes in the macromolecular structure dictates the biological activity and, consequently, how they can be explored in drug discovery. Most of the current studies on alkylphospholipids focuses on their physicochemical interactions with cholesterol and sphingolipids in lipid rafts that, because of the variability and complexity of the membrane phases, hardly can provide structural data in the X‐ray crystallography assays necessary for molecular modeling studies. Therefore, by exploring the inhibition of human cytidine triphosphate:phosphocholine cytidyltransferase as an alternative and, probably, complementary hypothesis to the membrane rafts, a helpful strategy can be provided to overcome the clinical limitations of alkylphospholipids.     

The Inhibitory Role of Rab11b in Osteoclastogenesis through Triggering Lysosome-Induced Degradation of c-Fms and RANK Surface Receptors

Rab11b, abundantly enriched in endocytic recycling compartments, is required for the establishment of the machinery of vesicle trafficking. Yet, no report has so far characterized the biological function of Rab11b in osteoclastogenesis. Using in vitro model of osteoclasts differentiated from murine macrophages like RAW-D cells or bone marrow-derived macrophages, we elucidated that Rab11b served as an inhibitory regulator of osteoclast differentiation sequentially via (i) abolishing surface abundance of RANK and c-Fms receptors; and (ii) attenuating nuclear factor of activated T-cells c1 (NFATc-1) upstream signaling cascades, following RANKL stimulation. Rab11b was localized in early and late endosomes, Golgi complex, and endoplasmic reticulum; moreover, its overexpression enlarged early and late endosomes. Upon inhibition of lysosomal function by a specific blocker, chloroquine (CLQ), we comprehensively clarified a novel function of lysosomes on mediating proteolytic degradation of c-Fms and RANK surface receptors, drastically ameliorated by Rab11b overexpression in RAW-D cell-derived osteoclasts. These findings highlight the key role of Rab11b as an inhibitor of osteoclastogenesis by directing the transport of c-Fms and RANK surface receptors to lysosomes for degradation via the axis of early endosomes-late endosomes-lysosomes, thereby contributing towards the systemic equilibrium of the bone resorption phase.     

Photoluminescence and scintillation properties of Eu2O3–BaO–Nb2O5–TeO2 glass and glass ceramics

Journal of Materials Science: Materials in Electronics - 1Eu2O3–3BaO–20Nb2O5–76TeO2 glass and the corresponding glass-ceramics were synthesized with the aim to investigate the...     

Generation and characterization of ultrafast white-light continuum in condensed media

The generation of white-light continuum by femtosecond laser pulses in transparent condensed media is investigated comprehensively with 262-, 393-, and 785-nm pump wavelengths. We find that the ratio of the medium's bandgap energy to the photon energy of the incident wavelength determines the amount of anti-Stokes broadening, independently of the pump wavelength and the medium's bandgap. It is also shown that, although the amount of anti-Stokes broadening is greater for the longer pump wavelength, the shorter pump wavelength is more advantageous for generating shorter-wavelength continua in the UV region. In addition, a self-induced change in polarization of the white-light continuum that is generated is observed in an isotropic material with a cubic crystal structure, such as CaF2 and LiF. After the investigation of polarization, the frequency chirp of the continuum is characterized by the Kerr-gate method with 70-fs temporal and 10-nm wavelength resolution.     

Supramolecular Nanostructures of Chiral Perylene Diimides with Amplified Chirality for High‐Performance Chiroptical Sensing

Chiral supramolecular nanostructures with optoelectronic functions are expected to play a central role in many scientific and technological fields but their practical use remains in its infancy. Here, this paper reports photoconductive chiral organic semiconductors (OSCs) based on perylene diimides with the highest electron mobility among the chiral OSCs and investigates the structure and optoelectronic properties of their homochiral and heterochiral supramolecular assemblies from bottom‐up self‐assembly. Owing to the well‐ordered supramolecular packing, the homochiral nanomaterials exhibit superior charge transport with significantly higher photoresponsivity and dissymmetry factor compared with those of their thin film and monomeric equivalents, which enables highly selective detection of circularly polarized light, for the first time, in visible spectral range. Interestingly, the heterochiral nanostructures assembled from co‐self‐assembly of racemic mixtures show extraordinary chiral self‐discrimination phenomenon, where opposite enantiomeric molecules are packed alternately into heterochiral architectures, leading to completely different optoelectrical performances. In addition, the crystal structures of homochiral and heterochiral nanostructures have first been studied by ab initio X‐ray powder diffraction analysis. These findings give insights into the structure–chiroptical property relationships of chiral supramolecular self‐assemblies and demonstrate the feasibility of supramolecular chirality for high‐performance chiroptical sensing.