Search results for "Electrophoresi"
showing 10 items of 1009 documents
Acidic cytosolic proteins are preferentially imported into rat liver lysosomes.
1998
Previous studies have reported that lysosomes isolated from human diploid fibroblasts and from rat liver can selectively import and degrade specific proteins. We have now reinvestigated this selectivity using an in vitro assay with rat liver lysosomes and an extract of cytosolic proteins prepared from cultured cells labeled to equilibriums with [35S-]methionine. Analysis by two-dimensional gel electrophoresis and autoradiography of the cytosolic proteins bound to the lysosomal membrane and imported into the lysosomes shows that when all cytosolic proteins are simultaneously present in the in vitro assay the lysosomal uptake also occurs in a specific manner. These findings suggest that isola…
Chiral separation of oxprenolol by affinity electrokinetic chromatography-partial filling technique using human serum albumin as chiral selector
2005
The intrinsic characteristics of capillary electrophoresis have made this technique a powerful tool in the chiral separation field. The present paper deals with the enantiomeric separation of oxprenolol enantiomers by affinity electrokinetic chromatography-partial filling technique using human serum albumin (HSA) as chiral selector. Several experimental conditions and variables affecting the separation such as pH, HSA concentration and plug length, background electrolyte concentration, temperature and voltage were studied. Baseline separation of oxprenolol enantiomers was obtained in less than 8 min under the following selected conditions: electrophoretic buffer composed of 50 mM Tris-(hydr…
Evaluation of enantioselective binding of fluoxetine to human serum albumin by ultrafiltration and CE - Experimental design and quality considerations
2012
Several pharmacokinetic processes are affected by enantioselectivity (ES). At the level of distribution, protein binding (PB) is one of the most important. The enantioselective binding of fluoxetine (FLX) to HSA has been evaluated in this work by ultrafiltration of FLX–HSA mixtures and chiral analysis of unbound fractions by EKC-CD. PB, affinity constants (K) and ES were obtained for both enantiomers of FLX. In order to improve the consistency of the estimations, the evaluation of affinity constants of each enantiomer was performed using two designs, one keeping constant the total concentration of protein and varying the total concentration of the enantiomers, and the other in the opposite …
Identification and isolation of the primary aggregation factor from the cell membrane of the sponge Geodia cydonium
1985
The primary aggregation factor (pAF) of sponge cells is a glycoprotein that is firmly associated with the cell membrane. Polyspecific antibodies (anti-GM) prepared from sera raised against membranes of cells from the siliceous sponge Geodia cydonium were found to inhibit initial aggregation of homologous cells. The inhibition of aggregation, caused by anti-GM was neutralized by pAF. The pAF had been successfully solubilized and enriched by affinity chromatography, gel filtration and density gradient centrifugation, if checked by polyacrylamide gel electrophoresis in the presence of urea. The Mr of the native pAF was approximately 40 000 as estimated by gel filtration; under denaturing condi…
Mesoscopic Simulations of Polyelectrolyte Electrophoresis in Nanochannels
2011
We present the results of mesoscopic dissipative particle dynamics (DPD) simulations of coupled electrohydrodynamic phenomena on the micro- and nanoscale. The effects of electroosmotic flow and slippage combined with polyelectrolyte electrophoresis are investigated in detail, taking full account of hydrodynamic and electrostatic interactions. Our numerical results are in excellent agreement with analytical calculations.
Experimental determination of effective charges in aqueous suspensions of colloidal spheres
2003
Abstract We determined the low frequency conductivity σ , the phase behaviour and the shear modulus G of colloidal fluids, respectively solids prepared from deionised aqueous suspensions of highly charged spherical particles. Conductivity measures the number of freely moving small ions Z * σ and thus relates to the ion condensation process in the electric double layer under conditions of finite macro-ion concentrations. Phase behaviour and elasticity data are consistently described by a Debye–Huckel pair potential assuming pair-wise additive macro-ion interactions. Like Z * σ , also the effective charges Z * G derived from the elasticity data scales with the ratio of macro-ion radius to Bje…
Interaction ofEscherichia colihemolysin with biological membranes
2001
Escherichia coli hemolysin (HlyA) is a membrane-permeabilizing protein belonging to the family of RTX-toxins. Lytic activity depends on binding of Ca2(+) to the C-terminus of the molecule. The N-terminus of HlyA harbors hydrophobic sequences that are believed to constitute the membrane-inserting domain. In this study, 13 HlyA cysteine-replacement mutants were constructed and labeled with the polarity-sensitive fluorescent probe 6-bromoacetyl-2-dimethylaminonaphthalene (badan). The fluorescence emission of the label was examined in soluble and membrane-bound toxin. Binding effected a major blue shift in the emission of six residues within the N-terminal hydrophobic domain, indicating inserti…
Strong Cooperativity and Loose Geometry between CUB Domains Are the Basis for Procollagen C-Proteinase Enhancer Activity
2009
Procollagen C-proteinase enhancers (PCPE-1 and -2) specifically activate bone morphogenetic protein-1 (BMP-1) and other members of the tolloid proteinase family during C-terminal processing of fibrillar collagen precursors. PCPEs consist of two CUB domains (CUB1 and CUB2) and one NTR domain separated by one short and one long linker. It was previously shown that PCPEs can strongly interact with procollagen molecules, but the exact mechanism by which they enhance BMP-1 activity remains largely unknown. Here, we used a series of deletion mutants of PCPE-1 and two chimeric constructs with repetitions of the same CUB domain to study the role of each domain and linker. Out of all the forms teste…
Organoplatinum(II) Complexes Self-Assemble and Recognize AT-Rich Duplex DNA Sequences
2021
The specific recognition of AT-rich DNA sequences opens up the door to promising diagnostic and/or therapeutic strategies against gene-related diseases. Here, we demonstrate that amphiphilic PtII complexes of the type [Pt(dmba)(N∧N)]NO3 (dmba = N,N-dimethylbenzylamine-κN, κC; N∧N = dpq (3), dppz (4), and dppn (5)) recognize AT-rich oligonucleotides over other types of DNA, RNA, and model proteins. The crystal structure of 4 shows the presence of significant π-stacking interactions and a distorted coordination sphere of the d8 PtII atom. Complex 5, containing the largest π-conjugated ligand, forms supramolecular assemblies at high concentrations under aqueous environment. However, its aggreg…
Application of Cell Culture for the Production of Bioactive Compounds from Sponges: Synthesis of Avarol by Primmorphs from Dysidea avara
2000
Among all metazoan phyla, sponges are known to produce the largest number of bioactive compounds. However, until now, only one compound, arabinofuranosyladenine, has been approved for application in humans. One major obstacle is the limited availability of larger quantities of defined sponge starting material. Recently, we introduced the in vitro culture of primmorphs from Suberites domuncula, which contain proliferating cells. Now we have established the primmorph culture also from the marine sponge Dysidea avara and demonstrate that this special form of sponge cell aggregates produces avarol, a sesquiterpenoid hydroquinone, known to display strong cytostatic activity especially against ma…