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At least 829 records · Page 46Linked to original sources
PubMed2006

The in-vitro bioactivity of mesoporous bioactive glasses.

Ordered mesoporous bioactive glasses (MBGs) with different compositions were prepared by using nonionic block copolymer surfactants as structure-directing agents through an evaporation-induced self-assembly process. Their in-vitro bioactivities were studied in detail by electron microscopy, Fourier-transform infrared spectroscopy, and inductively coupled plasma (ICP) atomic emission spectroscopy. The ICP element analysis results were further calculated in terms of the total consumption of Ca and P, Delta[Ca]/Delta[P] ratios, and ionic activity product (IP) of hydroxyapatite. Through the above analysis, it is clear that MBGs show a different structure-bioactivity correlation compared to conventional sol-gel-derived BGs. The in vitro bioactivity of MBGs is dependent on the Si/Ca ratio in the network when the other material parameters such as the mesostructure and texture properties (pore size, pore volume) are controlled. MBG 80S15C with relatively lower calcium content exhibits the best in vitro bioactivity, in contrast to conventional sol-gel-derived BGs where usually higher calcium percentage BGs (e.g. 60S35C) show better bioactivity. Calcination temperature is another important factor that influences the in vitro bioactivity. According to our results, MBGs calcined at 973K may possess the best in vitro bioactivity. The influences of the composition and calcination temperature upon bioactivity are explained in terms of the unique structures of MBGs.

Bone Substitutes↗
PubMed2006

Layer-by-layer assembly of cationic lipid and plasmid DNA onto gold surface for stent-assisted gene transfer.

Intravascular stent-assisted gene transfer is an advanced approach for the therapy of vascular diseases such as atherosclerosis and stenosis. This approach requires a stent that allows local and efficient administration of therapeutic genes to the target cells at the vascular wall. To create such a stent, a method was developed for loading plasmid DNA onto the metal surface. The method involves the formation of self-assembled monolayer on the noble metal surface followed by electrostatic layer-by-layer (LBL) assembly of a cationic lipid/plasmid DNA complex and free plasmid DNA. In this in vitro feasibility study, the thin plainer film and the wire of gold were used as a substrate. The LBL assembly process was characterized by surface plasmon resonance spectroscopy and static contact angle measurement. Plasmid DNA loaded in the multilayer exhibited improved resistance against nuclease digestion. When cultured directly on the DNA-loaded surface, cells were transfected to express exogenous gene in the DNA loading-dependent manner. Plasmid DNA could also be transferred to endothelial cells from its apical side by placing the DNA-loaded gold wire onto the cell layer.

Cations↗
PubMed2004

Immunoassays based on electrochemical detection using microelectrode arrays.

We show that CombiMatrix's VLSI arrays of individually addressable electrodes, using conventional CMOS integrated circuitry, can be used in detecting various analytes via immunoassay protocols. These microarrays provide over 1000 electrodes per square centimeter. The chips are coated with a porous material on which specific affinity tags are synthesized proximate to selected electrode sites. CombiMatrix microarrays are used to develop spatially multiplexed assay formats for biological entities over a wide range of sizes, from small molecules to cells. Antibodies are tagged with coded affinity labels and then allowed to self-assemble on the appropriate electrode assay sites. Each analyte-specific antibody is chaperoned to individual, predetermined locations by the self-assembly process. The resulting chip can perform numerous different analyte-specific immunoassays, simultaneously. We present new detection technologies based upon the use of the active individually addressable microelectrodes on the chip: redox enzyme amplified electrochemical detection. The results for human alpha1 acid glycoprotein, ricin, M13 phage, Bacillus globigii spores, and fluorescein indicate that this method is one of the most sensitive available, with limits of detection in the attomole range. The detection range is 4-5 logs of analyte concentration, with an assay volume of 50 microl or less. The system provides for a host of multiplexed immunoassays because of the large number of electrodes available. We show how the assays can be optimized for maximum performance on the CombiMatrix microarray platform.

Biosensing Techniques↗
PubMed2005

Site-specific polymer modification of therapeutic proteins.

Recent advances in chemoselective ligation technology have made possible the modification of proteins with polymers in a site-specific and controlled manner. These approaches rely on the incorporation of chemoselective anchors into the protein backbone by either chemical or recombinant means, and subsequent modification with a polymer carrying a complementary linker. As a result, the assembly process and the covalent structure of the resulting protein-polymer conjugate are completely controlled, enabling the rational optimization of drug properties, in particular efficacy and pharmacokinetic properties. Application of chemoselective ligation technologies to cytokines and chemokines has led to the generation of new lead proteins for use as erythropoietic agents and HIV fusion inhibitors.

Drug Design↗
PubMed2005

Identification of a multicomponent complex required for outer membrane biogenesis in Escherichia coli.

Gram-negative bacteria have an outer membrane (OM) that functions as a barrier to protect the cell from toxic compounds such as antibiotics and detergents. The OM is a highly asymmetric bilayer composed of phospholipids, glycolipids, and proteins. Assembly of this essential organelle occurs outside the cytoplasm in an environment that lacks obvious energy sources such as ATP, and the mechanisms involved are poorly understood. We describe the identification of a multiprotein complex required for the assembly of proteins in the OM of Escherichia coli. We also demonstrate genetic interactions between genes encoding components of this protein assembly complex and imp, which encodes a protein involved in the assembly of lipopolysaccharides (LPS) in the OM. These genetic interactions suggest a role for YfgL, one of the lipoprotein components of the protein assembly complex, in a homeostatic control mechanism that coordinates the overall OM assembly process.

Bacterial Outer Membrane Proteins↗
PubMed2004

A refined circular template matching method for classification of human cytomegalovirus capsids in TEM images.

An automatic image analysis method for describing, segmenting, and classifying human cytomegalovirus capsids in transmission electron micrograph (TEM) images of host cell nuclei has been developed. Three stages of the capsid assembly process in the host cell nucleus have been investigated. Each class is described by a radial density profile, which is the average grey-level at each radial distance from the center. A template, constructed from the profile, is used to find possible capsid locations by correlation based matching. The matching results are further refined by size and distortion analysis of each possible capsid, resulting in a final segmentation and classification.

Capsid↗
PubMed2005

Hydrophobins: proteins with potential.

Hydrophobins are self-assembling proteins of fungal origin. Their ability to self-assemble into an amphipathic membrane is of interest for many different applications, ranging from medical and technical coatings to the production of proteinaceous glue and cosmetics. Assembled hydrophobins can modify surface characteristics, thus controling the binding properties of the surface; for example, enzymes can be actively and non-covalently immobilized on electrode surfaces and medical coatings can be improved for biocompatibility. Over the past few years research on hydrophobins has contributed to a better understanding of the self-assembly process and is generating more handles to control and manipulate the process. This knowledge could have an immediate effect on production levels, which are not yet adequate, and provide the boost needed for hydrophobins to reach their full potential.

Biocompatible Materials↗
PubMed2005

Protein NO52--a constitutive nucleolar component sharing high sequence homologies to protein NO66.

The nucleolus is the most prominent intranuclear structure of almost all protein-synthesizing cells. It compromises a well-defined functional compartmentalization and a high complexity of molecular constituents. Here, we report on the identification and molecular characterization of a novel constitutive nucleolar component--protein NO52--that is present in diverse species from Xenopus laevis to human. The cDNA-deduced amino acid sequence of protein NO52 defines a polypeptide of a calculated mass of 52.8 kDa and an isoelectric point of 6.7. Inspection of the primary sequence disclosed that the protein contains a JmjC domain and is highly sequence-related to the recently described nucleolar protein NO66. Immunolocalization studies revealed that protein NO52 is highly concentrated in the granular component of nucleoli and this characteristic intranuclear distribution is significantly affected by treatment of cells with (i) RNase A, (ii) actinomycin D and (iii) serum starvation. Interestingly, protein NO52 has been identified as a constituent of free preribosomal particles but is absent from cytoplasmic ribosomes. Analyses of immunocomplexes isolated from cellular extracts with an NO52-specific antibody by MALDI mass spectrometry further confirmed the interaction of protein NO52 with various ribosomal proteins as well as with a distinct set of non-ribosomal nucleolar proteins. The dependence of the nucleolar accumulation of the protein on ongoing rRNA transcription and the cellular metabolic state strongly suggest that protein NO52 is directly involved in ribosome biogenesis, most likely during the assembly process of preribosomal particles.

Amino Acid Sequence↗
PubMed2005

Evaluation of a chylomicron flow blocking approach to investigate the intestinal lymphatic transport of lipophilic drugs.

The purpose of this study was to examine the feasibility of investigating the lymphatic transport of drugs in vivo utilizing known chylomicron flow blocking substances. Vitamin D(3) (0.5 mg/kg), a model lipophilic molecule, was administered to rats with blocked chylomicron flow, induced by either cycloheximide injection (3 mg/kg), colchicine injection (5 mg/kg) or intraduodenal infusion of pluronic L-81 (1 mg/h). The effect of these experimental models on the absorption of Vitamin D(3) was compared to the outcomes of the mesenteric lymph duct cannulated rat model. The oral d-xylose loading test was used to verify that other intestinal absorptive functions were not affected. Colchicine treatment induced severe adverse effects whereas pluronic L-81 and the cycloheximide models did not affect other absorption pathways and did not cause apparent adverse effects. Vitamin D(3) absorption in these two models was in good correlation to the mesenteric lymph duct cannulation model (25% non-lymphatic relative bioavailability) indicating that the incorporation of the lipophilic molecule into the chylomicron is an essential step in the cascade of lymphatic absorption. Moreover, the data suggest that the drug association with the chylomicron occurs at an early stage of its assembly process. The results also specify that lymphatic absorption and portal blood absorption are separate pathways that are not affected by each other. In conclusion, the chemical blockage of chylomicron flow provides a potential approach for lymphatic transport investigation, and may elucidate processes involving in the absorption of lipophilic compounds.

Animals↗
PubMed2006

Mass spectrometry of hydrogen/deuterium exchange in 70S ribosomal proteins from E. coli.

The 70S ribosome from Escherichia coli is a supermacro complex (MW: 2.7MDa) comprising three RNA molecules and more than 50 proteins. We have for the first time successfully analyzed the flexibility of 70S ribosomal proteins in solution by detecting the hydrogen/deuterium exchange with mass spectrometry. Based on the deuterium incorporation map of the X-ray structure obtained at the time of each exchange, we demonstrate the structure-flexibility-function relationship of ribosome focusing on the deuterium incorporation of the proteins binding ligands (tRNA, mRNA, and elongation factor) and the relation with structural assembly processes.

Deuterium Exchange Measurement↗
PubMed2004

Thermodynamic method for prediction of surfactant-modified oil droplet contact angle.

A model applying surfactant self-assembly theory and classical thermodynamics has been developed to aid in the prediction of solid surface cleaning by aqueous surfactant solutions. Information gained from a combination of surfactant self-assembly behavior and cleaning system parameters, such as oil species, surfactant type, temperature, alkalinity, and solid surface type has been shown to provide insight into surface cleaning. The model combines minimization of free energy, pertinent component distribution mechanisms, and surfactant self-assembly processes to provide a methodology for the predicting of oil droplet contact angles. Such predictive capabilities will allow for the development of beneficial environmental and economic changes to industrial and commercial surface cleaning and degreasing processes. Results from the model will be compared to experimental data to verify the capability of the theory to account for the effect of solutions parameters on oil droplet behavior. The model, while approximate in nature, has shown a remarkable quantitative predictive ability.

Journal Article↗
PubMed2006

Preparation of hollow capsule-stabilized gold nanoparticles through the encapsulation of the dendrimer.

The narrow-dispersed dendrimer-encapsulated gold nanoparticles without agglomeration were prepared from the grafted-dendrimers on the surface of the silica microspheres, successively by complexation of the gold chloride anion with the nitrogen atom of the dendrimer and the reduction with sodium borohydride as reductant. The hollow capsule-stabilized gold nanoparticles were prepared through the encapsulation of dendrimer successively by LBL self-assembly process of polyelectrolytes with modified-silica as template and removal of the silica core by hydrofluoric acid. The size of gold nanoparticles was 2.3+/-0.8 nm. The catalytic activity of the capsule-stabilized gold nanoparticles was investigated in the reduction reaction of 4-nitrophenol to 4-aminophenol with sodium borohydride as reductant.

Journal Article↗
PubMed2006

Polymer-mediated chain-like self-assembly of functionalized gold nanoparticles.

We report an easy solution phase template-based method to assemble mercaptoundecanoic acid-functionalized gold nanoparticles (MUA-GNPs) along poly(ethylene oxide) (PEO) chains. Transmission electron microscopy (TEM) images show one-dimensional and two-dimensional chain-like sequences of GNPs resembling PEO chains. The progress of the assembly was monitored by the evaluation of surface plasmon resonance band of MUA-GNPs with time by UV-vis spectroscopy. The assembly process is a result of hydrogen bonding interaction between the ethereal oxygen of PEO and carboxylic acid group of MUA attached to GNPs surface, which was confirmed through FTIR spectroscopy. The interaction between PEO and MUA-GNPs was further confirmed by thermal analysis using differential scanning calorimetry.

Journal Article↗
PubMed2006

Well-ordered mesoporous bioactive glasses (MBG): a promising bioactive drug delivery system.

The local drug release system is considered to be an alternative to treat the bone infection. In this paper, well-ordered mesoporous bioactive glasses (MBG) with high specific surface area have been synthesized in aqueous solution by a two-step acid-catalyzed self-assembly process combined with hydrothermal treatment. Gentamicin was encapsulated into the MBG by adsorption method and in vitro release of gentamicin from MBG was performed in distilled water and modified simulated body fluid (SBF), respectively. The results showed that the amount of drug loading of MBG was three times more than that of conventional sol-gel 58S. The outcomes of drug release in distilled water and in SBF showed that M58S effectively decreased the initial burst. During the release period, gentamicin was released from the M58S at a much lower release rate as compared to that from 58S after soaking in distilled water and SBF. Furthermore, the drug release was sensitive to the pH and ionic concentration of the release medium suggesting possible controls of the release rate. In addition, in contrast to conventional sol-gel 58S, M58S had higher ability to induce hydroxyapatite (HAp) formation. Therefore, well-ordered mesoporous bioactive glasses might be used as a bioactive drug release system for preparation of bone implant materials.

Drug Delivery Systems↗
PubMed2006

Development of a piezoelectric immunosensor for the measurement of paclitaxel.

This paper describes the development of a piezoelectric immunosensor for the measurement of paclitaxel (taxol), a natural anti-cancer agent. An antibody specific for taxanes was immobilized onto the surface of quartz crystals by means of the layer-by-layer self-assembly technique. The immobilization was achieved using electrostatic interactions between a precursor layer and the antibody molecules. The assembly process was monitored by a quartz crystal microbalance (QCM) and the topography of the modified quartz crystals was investigated by means of atomic force microscopy. The specific interaction of the immobilized antibody with paclitaxel in solution at different concentrations was monitored as a change in resonant frequency of the modified crystal. Moreover, the influence of non-specific adsorption was also characterized. The results show that the proposed immunosensor offers a promising alternative to classical analytical methods for a fast and easy determination of paclitaxel.

Animals↗
PubMed2004

Structure of the coat protein in Pf1 bacteriophage determined by solid-state NMR spectroscopy.

The atomic resolution structure of Pf1 coat protein determined by solid-state NMR spectroscopy of magnetically aligned filamentous bacteriophage particles in solution is compared to the structures previously determined by X-ray fiber and neutron diffraction, the structure of its membrane-bound form, and the structure of fd coat protein. These structural comparisons provide insights into several biological properties, differences between class I and class II filamentous bacteriophages, and the assembly process. The six N-terminal amino acid residues adopt an unusual "double hook" conformation on the outside of the bacteriophage particle. The solid-state NMR results indicate that at 30 degrees C, some of the coat protein subunits assume a single, fully structured conformation, and some have a few mobile residues that provide a break between two helical segments, in agreement with structural models from X-ray fiber and neutron diffraction, respectively. The atomic resolution structure determined by solid-state NMR for residues 7-14 and 18-46, which excludes the N-terminal double hook and the break between the helical segments, but encompasses more than 80% of the backbone including the distinct kink at residue 29, agrees with that determined by X-ray fiber diffraction with an RMSD value of 2.0 A. The symmetry and distance constraints determined by X-ray fiber and neutron diffraction enable the construction of an accurate model of the bacteriophage particle from the coordinates of the coat protein monomers.

Bacteriophage Pf1↗
PubMed2005

Towards a structural understanding of the fibrillization pathway in Machado-Joseph's disease: trapping early oligomers of non-expanded ataxin-3.

Machado-Joseph's disease is caused by a CAG trinucleotide repeat expansion that is translated into an abnormally long polyglutamine tract in the protein ataxin-3. Except for the polyglutamine region, proteins associated with polyglutamine diseases are unrelated, and for all of these diseases aggregates containing these proteins are the major components of the nuclear proteinaceous deposits found in the brain. Aggregates of the expanded proteins display amyloid-like morphological and biophysical properties. Human ataxin-3 containing a non-pathological number of glutamine residues (14Q), as well as its Caenorhabditis elegans (1Q) orthologue, showed a high tendency towards self-interaction and aggregation, under near-physiological conditions. In order to understand the discrete steps in the assembly process leading to ataxin-3 oligomerization, we have separated chromatographically high molecular mass oligomers as well as medium mass multimers of non-expanded ataxin-3. We show that: (a) oligomerization occurs independently of the poly(Q)-repeat and it is accompanied by an increase in beta-structure; and (b) the first intermediate in the oligomerization pathway is a Josephin domain-mediated dimer of ataxin-3. Furthermore, non-expanded ataxin-3 oligomers are recognized by a specific antibody that targets a conformational epitope present in soluble cytotoxic species found in the fibrillization pathway of expanded polyglutamine proteins and other amyloid-forming proteins. Imaging of the oligomeric forms of the non-pathological protein using electron microscopy reveals globular particles, as well as short chains of such particles that likely mimic the initial stages in the fibrillogenesis pathway occurring in the polyglutamine-expanded protein. Thus, they constitute potential targets for therapeutic approaches in Machado-Joseph's disease, as well as valuable diagnostic markers in disease settings.

Amino Acid Sequence↗
PubMed2006

Self-association of the transmembrane domain of an anthrax toxin receptor.

Protective antigen (PA), lethal factor (LF) and edema factor (EF) are secreted individually by Bacillus anthracis. These components of anthrax toxin must then assemble into complexes to intoxicate mammalian cells. Toxin assembly initiates when molecules of PA bind mammalian receptors ANTXR1/2 and are cleaved by surface proteases into 20 kDa and 63 kDa fragments. After PA20 dissociates, receptor-bound PA63 homo-oligomerizes into heptamers. Oligomeric PA63 binds EF and LF and these complexes are internalized into an acidic compartment where the two enzymatic components are translocated across the membrane by a channel formed by heptameric PA63. Since oligomerization of PA63 is required to bind and translocate the enzymatic components, we sought to determine whether interactions between toxin receptors could facilitate the assembly process. In the present work, we performed a co-immunoprecipitation experiment to demonstrate that ANTXR1 is oligomeric in mammalian cells. Computer modeling predicted the self-association of the ANTXR1 transmembrane domain and we detected oligomerization of ANTXR1 transmembrane domain peptides in the membrane-mimetic environment of SDS micelles using fluorescence resonance energy transfer. Furthermore, the ANTXR1 transmembrane domain mediated oligomerization of a reporter protein construct in a bacterial membrane. In both assays, mutations that disrupted the interaction were consistent with the interaction being mediated through an asymmetric binding interface. Mutations that impaired self-association of the transmembrane domain reduced the rate of PA63 heptamer formation on the mammalian cell surface. Our findings indicate that ANTXR1 transmembrane domains self-associate and that these interactions may stabilize intermediate oligomerization states of ANTXR1-PA63 complexes.

Amino Acid Sequence↗