Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound   Target Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound

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Query: EC:3.2.1.17 (lysozyme)
21,489 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The thermodynamic constants, associated with the interaction of three proteins with triazine dye affinity sorbents, have been derived from bath and frontal analysis experiments. In cases where mass-transfer restrictions are very high, calculation of the thermodynamic constants directly from frontal analysis experiments could not be achieved. In such cases, a portion of the adsorbate was always present in the effluent, a situation which has its effect as the split peak phenomenon. With Fractogel-based triazine dye affinity sorbents none of the test proteins applied in frontal analysis were adsorbed. A similar behaviour was observed for a Cellufine sorbent during the adsorption of human serum albumin and the Blue Sepharose CL6B sorbent during the adsorption of alcohol dehydrogenase, which displayed much slower apparent adsorption kinetics than observed in the bath experiments. These phenomena were shown to be associated with changes in the gel structure, caused in part by the column packing procedure. Silica-based sorbents performed better in the adsorption of lysozyme in the column mode than soft-gel affinity sorbents, as was evident in the higher capacities and steeper breakthrough curves. At high protein concentrations (feedstock concentration greater than 0.2 mg/ml) breakthrough curves obtained with small- and large-particle-size sorbents, but of constant pore size, were found to be identical. This finding demonstrates that the use of small-particle-size sorbents (e.g. particle diameter, dp less than or equal to 5 microns) for the preparative isolation of proteins may not be justified when operating in the overload mode. With other higher-molecular-weight proteins and the silica-based sorbent systems examined, the small-particle-size sorbents (dp = 5 microns) displayed less symmetrical shapes of their breakthrough curves than the larger-particle-size and soft-gel sorbents. This behaviour was further exacerbated when non-porous glass or silica-based sorbents were utilized. These non-porous affinity sorbents displayed nearly rectangular breakthrough shapes at the onset of the adsorption process, but comparatively slow adsorption kinetics became evident as saturation was approached. This phenomenon has been attributed to surface rearrangement and/or reorientation of the adsorbed proteins, particularly with sorbents of high ligand densities.
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PMID:High-performance liquid chromatography of amino acids, peptides and proteins. XCV. Thermodynamic and kinetic investigations on rigid and soft affinity gels with varying particle and pore sizes: comparison of thermodynamic parameters and the adsorption behaviour of proteins evaluated from bath and frontal analysis experiments. 215 23

Macromolecular adsorption is known to occur as a complex process, often in a series of steps. Several models are discussed in the literature which describe the microscopic structure of the adsorbate. In the present study we investigated the adsorption of hen egg white lysozyme on alkylated silicon oxide surfaces. A combination of fluorescence excitation in the evanescent field and fluorescence recovery after photobleaching allowed us to measure the amount of adsorbed fluorescent lysozyme and the equilibrium exchange kinetics with molecules in solution. We found that a model with at least three classes of adsorbed molecules is necessary to describe the experimental results. A first layer is formed by the molecules which adsorb within a short time after the beginning of the incubation. These molecules make up approximately 65% of the final coverage. They are quasi-irreversibly adsorbed and do not measurably exchange with bulk molecules within one day even at temperatures up to 55 degrees C. A second layer, which reaches equilibrium only after several hours of incubation, shows a pronounced exchange with bulk molecules. The on-off kinetics show a distinct temperature dependence from which an activation barrier of delta E approximately 22 kcal/mol is derived. A third layer of molecules that exchange rapidly with the bulk can be seen to comprise approximately 10% of the total coverage. The exchange rate is on the order of fractions of a second. The binding of the latter two classes of adsorbed molecules is exothermic. From the temperature dependence of the coverage, the binding enthalpy of the slowly exchanging layer was estimated to be delta Hads approximately 3.8 kcal/mol. The second and third class of molecules remain enzymatically active as a muramidase, which was tested by the lysis of the cell walls of Micrococcus lysodeiktikus. The molecules in the first layer, on the other hand, showed no enzymatic activity.
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PMID:Multilayer adsorption of lysozyme on a hydrophobic substrate. 230 2

Foamy alveolar macrophages (FAM) are observed in lungs injured by Bleomycin (BLM), but their relation to pulmonary fibrosis is not clearly understood. We purified FAM from BLM-instilled rat lungs by density gradient centrifugation on Percoll, and studied the effect of FAM on pulmonary fibrosis. The cells lavaged from the rat lungs 14 days after the administration of BLM (B) or saline (S), were applied on Percoll. After centrifugation, the cells layered on each interface were collected and named as SI, SII, SIII, and BI, BII, BIII in order of gravity. The BI layer included 8.5% of unfractionated cells (U). These BI cells were viable (88%), significantly larger than the others, nonspecific esterase positive cells, and included much ferritin and lysozyme, and were morphologically identified as alveolar macrophages (AM). Therefore, we called the BI cells FAM. We estimated the capacity of FAM (2.5 X 10(5] to synthesize DNA (3H-thymidine uptake) and RNA (3H-uridine uptake), and the activities of silica-stimulated FAM to cause proliferation of mouse thymocytes (IL-1 activity) and rat lung fibroblasts (FP activity), and to produce PGE2. FAM has a lower mitogenic activity but did not have been protein synthetic activity as compared with the others. Silica-stimulated FAM released less IL-1 than BII or BIII, and induced less fibroblast growth than BII, but induced as much as BIII, possibly because of the increased capacity of BIII cells to produce PGE2, which is known to inhibit fibroblast growth. In this way, FAM were considered to be "already activated" rather than "highly activated" cells, but the presence of FAM suggested that smaller or denser AM might receive bleomycin stimulation and release fibrogenic mediators (IL-1 or MDGF) into the alveolar spaces during FAM formation, and that AM might participate in the fibrogenic responses.
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PMID:[The effect of foamy alveolar macrophages presented in bleomycin-injured rat lungs in pulmonary fibrosis]. 247 35

Male Sprague-Dawley rats were subjected to a single, intratracheal instillation of 30 mg Min-U-Sil silica in sterile saline and were sacrificed 3, 7, or 14 days following instillation. Control animals were instilled with sterile saline only. Silica instillation produced an inflammatory reaction followed by histological changes characteristic of lung fibrosis. Thickened alveolar septa associated with inflammatory cells transforming into large multifocal fibrotic nodules were detected in silica-exposed animals. Increased numbers of bronchoalveolar cells (principally macrophages), elevated levels of protein (principally serum albumin), and lysozyme, proteolytic (trypsin-like), and myeloperoxidase activities were detected in lavage fluids obtained from animals instilled with silica. These factors (except for lysozyme activity) were elevated above control levels from 3 to 7 days postinstillation and declined to near control levels by Day 14. The rate of DNA, collagen, and noncollagen protein synthesis was significantly elevated in lung tissue minces from silica-treated rats 3 and 7 days after instillation. Elevated levels of total protein, and lung collagen in particular, were observed 9 weeks after insult. Lavage fluid from silica-instilled rats stimulates DNA synthesis in cultures of proliferating and quiescent rat lung fibroblasts. Lavage fluid from silica-instilled rats also stimulates lung fibroblasts to increase collagen and noncollagen protein synthesis.
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PMID:Regulation of lung fibroblast proliferation and protein synthesis by bronchiolar lavage in experimental silicosis. 301 63

Deposits on soft contact lenses of high water content were investigated morphologically and chemically and compared with those on conventional soft contact lenses of poly (2-hydroxyethyl methacrylate). The material of the lenses examined in this investigation was the crosslinked copolymer of methyl methacrylate and N-vinylpyrrolidone with a water content higher than 70%. Morphologically, the deposits on the lenses with high water content were found to have no characteristics distinguishable from those on conventional lenses. By the electron microscopic observation of the cross section of a lens that had become opaque, it was confirmed that the deposit was on the lens surface and that no deposit was within the lens. Some spots on the lenses were recognized as colonies of microorganisms, but the majority of the spots had no involvement by microorganisms. Surface analysis with Fourier transform infrared spectrometer (FT-IR) confirmed that the main component of the filmy deposit was protein. Protein was detected in most of the deposits. The amino acid compositions of the proteins were found to be close to that of lysozyme. From the elemental analysis of several spots, silicon, aluminum, iron, and some other elements were detected. The structural analysis of some spots by a laser Raman microprobe (MOLE) revealed the existence of lipids. In several cases, the deposits were found to have grown around a defect of the lens surface. A mechanism for the formation of deposits is suggested.
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PMID:Analysis of deposits on high water content contact lenses. 684 67

The preparation and characterization of high-surface-area polymeric substrates suitable for the microcalorimetry of protein adsorption are described. High-surface-area polystyrene, poly(styrene-co-butyl methacrylate) and poly(styrene-co-allyl alcohol) were prepared by adsorbing polymer from solution onto fumed silica. Verification of adsorption of polystyrene by silica was determined by noting peak shifts of the surface silanol group in the infrared. The amount of polymer adsorbed was determined from adsorption isotherms. The minimum thickness of polystyrene required to mask silicon oxide properties was found to be that thickness at which contact angles became constant, about 35 A. Polymer densities were measured. Water contact angles on each polymer surface indicate that poly(styrene-co-allyl alcohol) has the surface most wettable by water. Polymer-water interfacial energies were estimated from pendant drop results and a harmonic mean equation along with contact angles. Two methods were used to estimate the polar and dispersion components of the three polymers. Both methods predicted polystyrene to have the highest interfacial energy against water, and one method predicted poly(styrene-co-allyl alcohol) to have the lowest. A Wilhelmy plate study verified the change in interfacial properties as a function of contact time with water. A study of the heats of adsorption of lysozyme by each substrate using a modified Tien-Calvet microcalorimeter demonstrated the suitability of the substrates for microcalorimetry.
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PMID:Preparation and characterization of high-surface-area polymer substrates for microcalorimetry. 873 Nov 49

A new measuring method is described for obtaining a proton titration curve. The curve is obtained from a microporous composite membrane, consisting of polystyrene beads in an agarose matrix, with lysozyme molecules adsorbed to the bead surface. The membrane is incorporated into a sensor system by deposition on a silicon chip with a pH-sensitive ion-sensitive field effect transistor (ISFET) located in the middle of a Ag/AgCl electrode. The actual measurement is performed by creating a stepwise change in the salt concentration of the bathing electrolyte (the ion step) and measuring the ISFET potential versus the Ag/AgCl electrode. This potential shows a transient change in the ion step, which indicates a transient pH change in the membrane. This procedure is repeated at a series of pH values. Equations are presented to calculate the proton titration curve of the membrane from the amplitude and duration of the measured transients. Measurements show qualitative agreement between the curves obtained and equilibrium titration experiments on the same system.
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PMID:Constructing a Proton Titration Curve from Ion-Step Measurements, Applied to a Membrane with Adsorbed Protein 924 24

We have studied the adsorption of lysozyme layers at a hydrophobic silicon water interface using specular neutron reflection. The hydrophobic surface was obtained by self-assembly of a densely packed monolayer of octadecyltrichlorosilane (OTS) onto the natural silica layer on the smooth surface of a (111) silicon block. The effect of pH on the adsorbed lysozyme layer was examined at a constant lysozyme concentration of 0.03 g dm-3 and at a constant ionic strength of 0.02 M. Reflectivity profiles at different pH show that adsorption is irreversible with respect to pH, the composition and structure of the final layer being dependent on the route by which the pH was achieved. The adsorbed protein layer was found to divide into approximately two regions, a densely packed thin layer next to the OTS surface and a diffuse thicker layer extending into the bulk solution. None of the dimensions of this structure corresponds to those of the globular protein in solution, suggesting that, unlike its adsorption at the hydrophilic silica/water interface, lysozyme is denatured at the OTS/water surface. The irreversible adsorption is explained by the combined interaction of the hydrophobic attraction of the hydrophobic fragments in lysozyme to the OTS surface and electrostatic repulsion within the adsorbed layer. The hydrophobic surface induces the exposure of hydrophobic fragments from the lysozyme assembly. The thickness of the dense layer suggests that the denatured protein adsorbs in the form of peptide chains with the hydrophobic amino acid side chains attached to the OTS surface with the hydrophilic side chains extending into the bulk solution. Since lysozyme is more stable at pH 7 than at pH 4, the difference in initial adsorption is dominated by the greater relative stability of lysozyme to denaturation at the higher pH. A change of pH from 7 to 4 reduces the stability of the protein to unfolding and results in more adsorption than when the pH is changed in the opposite direction. Solution pH also affects the net charges within the hydrophilic tail region and the structural distribution of the tail region was found to vary with pH. Copyright 1998 Academic Press.
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PMID:The Denaturation of Lysozyme Layers Adsorbed at the Hydrophobic Solid/Liquid Surface Studied by Neutron Reflection. 976 46

This paper presents a picoliter sample preparation technique utilizing the flow-through principle, allowing on-line coupling of chromatographic systems to be made. The work was performed in order to investigate the characteristics and the physicochemical properties of the sample preparation using typical mobile phase conditions from mu-CLC (column liquid chromatography) separations. The device presented here is a pressure pulse-driven dispenser, formed by two silicon structures processed by conventional micromachining. The pressure pulse is generated in the flow-through channel by a piezoceramic element. Depending on the orifice size, the droplets ejected range between 30 and 200 pL. The maximum ejection frequency is 500 Hz, limited by resonances within the unit. A pyramid-shaped nozzle improves the directivity of the droplets since it reduces the wetting of the orifice front surface area. The risk of particles sticking close to the orifice is also minimized. The analyses of the deposited sample spots were carried out on a matrix-assisted laser desorption/ionization time-of-flight mass spectrometer with delayed extraction. It was possible to detect attomole amounts (159-248 amol) of various proteins (cytochrome c, ribonuclease A, lysozyme, and myoglobin) from a single droplet of matrix:analyte 1:1 (drop volume approximately 110 pL). Additionally, it was found that sample enrichment could be carried out using multiple depositions on the same spot; i.e., 31 nM of insulin was easily detected when more than four depositions were made on the same spot, while no detection was possible without sample enrichment. Size optimization of the MALDI sample spot gave target zones of 100-500-micron diameter that matched the size of the laser focal point and resulted in a considerably increased sample throughput.
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PMID:Picoliter sample preparation in MALDI-TOF MS using a micromachined silicon flow-through dispenser. 984 71

Coating hydrogel polymers onto solid substrates can reduce the adsorption of proteins onto these surfaces, but the extent of the reduction in protein adsorption is strongly dependent on how the surface layer is coated. We have examined the effect of coating conditions on the structure of thin polymer films formed from a number of poly(methacrylate)-based hydrogel polymers via the dip-coating method. We show in this work how the polarity of the solvent, the speed of lifting, and the annealing temperature affect the thickness and uniformity of ultrathin phosphorylcholine (PC)-incorporated polymer films coated on the surface of native oxide on silicon and the subsequent interaction of these coated surfaces with lysozyme molecules. Our results show that the uniformity of the polymer film, and thus the smoothness of the outer film surface, influence the extent of reduction in protein adsorption. We suggest that the reduction in lysozyme adsorption is the result of a layer of PC groups on the surface of the polymer film. The improvement of the smoothness of the film results in the formation of a close-packed PC layer on the outer surface of the polymer film, leaving few defects or cavities on which protein molecules can bind.
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PMID:The reduced adsorption of lysozyme at the phosphorylcholine incorporated polymer/aqueous solution interface studied by spectroscopic ellipsometry. 1045 63


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