Showing papers on "Bovine serum albumin published in 1999"
TL;DR: In this article, the reversibility, specificity, stability, and scaling of signal response to analyte mass were quantified for a porous silicon-based optical interferometric biosensor.
Abstract: The reversibility, specificity, stability, and scaling of signal response to analyte mass were quantified for a porous silicon-based optical interferometric biosensor. The sensor system studied consisted of a thin layer (5μm) of porous silicon modified with Protein A. The system was probed with various fragments of an aqueous Human IgG analyte. The sensor operates by measurement of the Fabry−Perot fringes in the white light reflection spectrum from the porous silicon layer. Molecular binding is detected as a shift in wavelength of these fringes. IgG was added to and removed from the protein A-modified surface by changing solution pH in a flow cell, and the system was found to be reversible through several on−off cycles. The molecule used to link protein A to the porous Si surface incorporated bovine serum albumin (BSA). This approach was found to completely eliminate signal due to nonspecific binding, tested by exposure of the sensor to the F(ab‘)2 fragment of IgG (which does not bind to protein A). The l...
453 citations
TL;DR: The results suggest that clusterin may play a sHSP-like role in cytoprotection, and at physiological concentrations, clusterin potently protected glutathione S-transferase and catalase from heat-induced precipitation and α-lactalbumin and bovine serum albumin from precipitation induced by reduction with dithiothreitol.
435 citations
TL;DR: A triple mutant constructed in this study exhibited no hemagglutination using sheep erythrocytes or hemoglobin binding activity, as determined by a solid-phase binding assay with horseradish peroxidase-conjugated human hemoglobin, indicating that the adhesin domains seem to be particularly important for P. gingivalis cells to agglutinate ery Throcytes and bind Hemoglobin, leading to heme acquisition.
319 citations
TL;DR: It is proposed that, considering the poor glucose control found in diabetics as well as the key role of oxidative stress in vascular complications, glycation‐mediated and free radical‐induced impairment of the antioxidant properties of albumin might be important parameters in vascular complication encountered in diabetes.
Abstract: Epidemiological data consistently show that reduced levels of serum albumin, which is the most abundant protein in plasma, are associated with an increased mortality risk. Various biological properties evidenced by direct effects of the albumin molecule may explain its beneficial effects. The present work aimed to investigate in vitro whether glycation or free radicals or both factors would affect the antioxidant properties of bovine serum albumin (BSA). Glycation was performed by long-term incubations (60 days) of BSA with increasing concentrations of glucose (up to 500 mmol/l) at 37°C. Minimally oxidized BSA was obtained after controlled incubations of dialyzed BSA samples with a water-soluble free radical generator [2,2′ azo-bis(2-amidinopropane) HCl]. The glycation-mediated modifications and the free radical-induced conformational changes of BSA were monitored using intrinsic fluorescence measurements of the tryptophan residues and acrylamide as a quenching agent. Thiol groups, Amadori glycophore cont...
301 citations
TL;DR: In this article, photolysis using ultraviolet light and graft polymerization of hydrophilic monomers onto the membrane surface was used to create more hyrophilic and lower fouling membrane surfaces.
273 citations
TL;DR: Steadystate and dynamic fluorescence measurements were used to investigate the interactions and structures of complexes formed between bovine serum albumin (BSA) and anionic, cationic, and nonioni as discussed by the authors.
Abstract: Steady-state and dynamic fluorescence measurements were used to investigate the interactions and structures of complexes formed between bovine serum albumin (BSA) and anionic, cationic, and nonioni ...
258 citations
TL;DR: In this article, an initial rate approach was used to study the reaction of peroxynitrite with human serum albumin (HSA) through stopped-flow spectrophotometry.
250 citations
TL;DR: It is concluded that the activation of NFkappaB/Rel proteins modulates chemokine production in PTC in response to albumin and is likely to have an important role in the mediation of tubulointerstitial injury in proteinuric renal disease.
Abstract: The transcription and translation of monocyte chemoattractant protein-1 (MCP-1), a CC chemokine, are increased in proximal tubule epithelial cells (PTC) stimulated with pathophysiologically relevant concentrations of albumin. The purpose of this study was to investigate whether nuclear factor kappaB (NFkappaB)/Rel proteins play a role in albumin-induced MCP-1 transcription. Confluent monolayers of rat PTC in primary culture were stimulated with delipidated bovine serum albumin. NFkappaB, the NFkappaB inhibitory protein (IkappaB), and MCP-1 transcription were assessed using electrophoretic mobility shift assays, Western immunoblotting, semiquantitative reverse transcription-PCR, and ribonuclease protection assays. Activation of NFkappaB by delipidated bovine serum albumin (15 mg/ml) was detectable within 2 h, maximal after 8 h, and maintained for at least 16 h of continuous exposure. Supershift analysis showed that the activated proteins were composed of p50/p50, p50/p65, and p50/c-Rel dimers. dimers. Cytoplasmic IkappaBalpha levels were decreased 30 min after stimulation and returned to unstimulated levels by 4 to 8 h. IkappaBbeta levels were decreased at 2 h and there was no recovery until 8 h. Inhibition of NFkappaB with pharmacologic agents (N-tosyl-phenylalanine chloromethyl ketone and dexamethasone) and an antisense oligonucleotide to the rat p65 subunit of NFkappaB significantly reduced MCP-1 transcription. The 3.6-kb 5' flanking region of the rat MCP-1 gene was cloned and sequenced, and two putative kappaB binding sites were identified within the enhancer region. Therefore, albumin increased NFkappaB and reduced IkappaB levels in PTC, and MCP-1 expression was dependent on NFkappaB activation. It is concluded that the activation of NFkappaB/Rel proteins modulates chemokine production in PTC in response to albumin and is likely to have an important role in the mediation of tubulointerstitial injury in proteinuric renal disease.
204 citations
TL;DR: It is reported that E2-BSA conjugate preparations, but not unconjugated E2, activate extracellular signal-regulated protein kinases (ERK1 and ERK2) in the SK-N-SH neuroblastoma cell line, raising concerns regarding the use of these reagents as E2 mimics.
Abstract: The steroid 17beta-estradiol (E2) acts to modulate transcription through classical nuclear estrogen receptors (ER-alpha and ER-beta). However, E2 also induces a number of rapid responses (<10 min) within cells, including cells devoid of classical ERs, consistent with the presence of a membrane receptor for E2. Membrane impermeable steroids, typically bovine serum albumin (BSA) conjugates, are commonly used to characterize these non-genomic actions of E2 to exclude the involvement of nuclear ERs. Here we report that E2-BSA conjugate preparations, but not unconjugated E2, activate extracellular signal-regulated protein kinases (ERK1 and ERK2) in the SK-N-SH neuroblastoma cell line, raising concerns regarding the use of these reagents as E2 mimics. Freshly prepared solutions of E2-BSA were found to contain free immunoassayable E2 (iE2), which could be removed by filtration. E2-BSA solutions devoid of free iE2 failed to compete for binding of 125I16alpha-iodo-E2 to ER-alpha or ER-beta. Furthermore, in contrast to E2, E2-BSA conjugates did not bind to ER-alpha or ER-beta as assessed by electrophoretic mobility shift analyses. Protein analysis demonstrated that certain E2-BSA preparations were of very high molecular weight, suggesting extreme protein cross-linking. These findings suggest that E2-BSA does not mimic E2 and is not an appropriate ligand for investigating estrogen receptors. This underscores the need to design stable, cell impermeable analogs of estrogen for the characterization of membrane estrogen receptors.
199 citations
TL;DR: This study substantiated that an adequate formulation could overcome denaturing effects of the methylene chloride/water interface upon a protein of interest to be encapsulated into microspheres.
193 citations
TL;DR: Polyacrylamide gel electrophoresis (PAGE) and isoelectric focusing (IEF) analyses of BSA released from these particles confirmed that the entrapped protein seemed to remain unaltered by the protein encapsulation process.
TL;DR: An immunochemical technique for the quantification of carbonyl groups in protein samples prepared from small tissue samples and cell cultures was developed and results were highly correlated with those of the standard DNPH-based spectrophotometric technique.
TL;DR: The potential applications of magnetic particles in several biomedical and biotechnology fields are discussed and the binding of the proteins and enzymes to magnetic particles was confirmed.
TL;DR: It is demonstrated that M6P‐modified bovine serum albumin (BSA) accumulates in slices of normal and cirrhotic human livers and can be applied as a selective drug carrier for HSC.
TL;DR: This work has combined the presented role of Cu2+ in S-nitrosothiol formation with the known destabilizing effect of Cu+, providing a unique simple picture where the redox state of copper determines either the NO release fromS-nitrogenothiols or the NO scavenging by thiol groups.
TL;DR: It is proposed that embryo culture media should contain both serum albumin and hyaluronan, while the transfer medium need only contain hyalonsonan, as the highest cell numbers and hatching rates obtained in this study occurred when both serum Albumin and Hyaluronans were present in the same medium.
Abstract: The effect of macromolecules on mouse embryo development and viability after culture in sequential media was investigated. It was found that high rates of viable blastocysts could be obtained in the absence of any macromolecule. Blastocyst cell numbers were increased when bovine serum albumin was present in the culture medium, although this benefit was not manifest after blastocyst transfer. Rather, the highest rates of implantation and fetal development after blastocyst transfer were observed when hyaluronan was the macromolecule in the culture media. Subsequent analysis revealed that the beneficial effects of hyaluronan were due to its presence in the transfer medium. As the highest cell numbers and hatching rates obtained in this study occurred when both serum albumin and hyaluronan were present in the same medium, it is proposed that embryo culture media should contain both serum albumin and hyaluronan, while the transfer medium need only contain hyaluronan.
TL;DR: The present study provides the possibility to achieve a long-term effective release of biologically active proteins from a Ca-P-coated metallic implant.
Abstract: Calcium phosphate (Ca-P) and bovine serum albumin (BSA) were coprecipitated as a coating on commercially pure titanium (cpTi) with a high protein loading (15 wt %) by employing a recently developed wet-chemistry technique. It was observed that the incorporation of BSA significantly modified the morphology, composition, and crystallinity of the Ca-P coating. The Ca-P coating without BSA is a mixture of hydroxyapatite (HA) and octacalcium phosphate (OCP) with sharp-edged thin OCP crystal plates on the top layer, whereas only an HA phase was detected in the Ca-P/BSA coating. The crystal plates in the latter had a more rounded appearance. The Ca-P/BSA coatings were immersed respectively in neutral (pH 7.4) and acidic (starting pH 4.0) phosphate-buffered saline (PBS) at 37°C over a 14-day period. No protein release was detected in the neutral PBS during the immersion; however, a continuous release of BSA was measured in the acidic PBS, subsequently leading to the formation of a very dense and well-adherent composite coating of BSA and Ca-P on cpTi. The present study provides the possibility to achieve a long-term effective release of biologically active proteins from a Ca-P-coated metallic implant. © 1999 John Wiley & Sons, Inc. J Biomed Mater Res, 46, 245–252, 1999.
TL;DR: In this paper, bovine serum albumin (BSA) was subjected to high pressure processing at low ionic strength and neutral pH and showed a reduction in protein surface hydrophobicity.
TL;DR: This chapter discusses the measurement of protein carbonyl by enzyme-linked immunosorbent assay, a colorimetric procedure that measures binding of dinitrophenylhydrazine (DNP).
Abstract: Publisher Summary This chapter discusses the measurement of protein carbonyl by enzyme-linked immunosorbent assay. Protein carbonyls are formed by a variety of oxidative mechanisms and are sensitive indices of oxidative injury. The conventional assay for protein carbonyls is a colorimetric procedure that measures binding of dinitrophenylhydrazine (DNP). Protein-bound DNP can also be measured, with increased sensitivity, either by HPLC (high-performance liquid chromatography), or using an anti-DNP antibody with Western blots or tissue sections. Samples containing protein are reacted with DNP then the protein is nonspecifically adsorbed to an ELISA plate. Unconjugated DNP and nonprotein constituents are easily washed away and give minimal interference. The adsorbed protein is probed with a commercial biotinylated anti-DNP antibody followed by streptavidin-linked horseradish peroxidase. Absorbances are related to a standard curve prepared for bovine serum albumin (BSA) containing increasing proportions of hypochlorous acid 1(HOCl)-oxidized protein that is calibrated colorimetrically.
Patent•
15 Jun 1999
TL;DR: Erythropoietin analog-human serum albumin (EPOa-hSA) fusion protein and methods of making and using the fusion protein were discussed in this article.
Abstract: Erythropoietin analog-human serum albumin (EPOa-hSA) fusion protein and methods of making and using the fusion protein.
TL;DR: The isolation and identification of a 23 kDa membrane protein from digitonin-solubilized rat brain mitochondrial fractions that binds 17beta-estradiol conjugated to bovine serum albumin at C-6 position (17beta-E-6-BSA), a ligand that also specifically binds nER is reported.
TL;DR: Subsequent reactions of the Amadori and Heyns rearrangement products, cross-linking, development of Maillard fluorescence, oxidation, and fragmentation, indicated that the alpha-hydroxy carbonyl group of Amador i products is more reactive than the aldehydo group of Heyni products.
Abstract: Glycation of bovine serum albumin by d-glucose and d-fructose under dry-heating conditions was studied. The reactivities of d-glucose and d-fructose, with respect to their ability to utilize primary amino groups of proteins, to cross-link proteins, to develop Maillard fluorescence, and to reduce protein solubility in the presence and absence of air (molecular oxygen) were investigated. d-Glucose showed a higher initial rate of utilization of primary amino groups than d-fructose, both in the presence and in the absence of oxygen. Subsequent reactions of the Amadori and Heyns rearrangement products, cross-linking, development of Maillard fluorescence, oxidation, and fragmentation, indicated that the α-hydroxy carbonyl group of Amadori products is more reactive than the aldehydo group of Heyns products. d-Fructose showed a greater sensitivity than d-glucose toward the presence of oxygen at the initial stages of the Maillard reaction. The presence or absence of oxygen in the glycation mixture did not seem to ...
TL;DR: Naturally occurring peptides derived from the major whey proteins, i.e. alpha-lactalbumin and beta- lactoglobulin in addition to bovine serum albumin, inhibit ACE and may represent nutraceutical/functional food ingredients for the prevention/treatment of high blood pressure.
Abstract: Angiotensin-I-converting enzyme (ACE) has been classically associated with the renin-angiotensin system which regulates peripheral blood pressure. Peptides derived from the major whey proteins, i.e. alpha-lactalbumin (alpha-la) and beta-lactoglobulin (beta-lg) in addition to bovine serum albumin (BSA), inhibit ACE. Some of these inhibitory peptides, i.e. alpha-lactorphin (alpha-la f(50-53)), beta-lactorphin (beta-lg f(102-105)), beta-lactotensin (beta-lg f(146-149) and albutensin A (BSA f(208-216)), have other bioactivities. The most potent lactokinin reported to date, (beta-lg f(142-148)), has an ACE IC50 of 42.6 mumol/l. While they do not have the inhibitory potency of synthetic drugs commonly used in the treatment of hypertension, these naturally occurring peptides may represent nutraceutical/functional food ingredients for the prevention/treatment of high blood pressure. Studies with gastric and pancreatic proteinase digests of whey proteins indicate that enzyme specificity rather than extent of hydrolysis dictates the ACE inhibitory potency of whey hydrolysates.
TL;DR: A novel Förster donor-acceptor dye pair for an immunoassay based on resonance energy transfer (RET) is characterized with respect to its photophysical properties and the applicability of this dye pair is demonstrated in a homogeneous competitive immunoASSay for the pesticide simazine.
TL;DR: In this paper, the adsorption of bovine serum albumin and HSA at the air/water interface has been studied by neutron specular reflection, and the effect of bulk protein concentration on the adorbed amount and the total thickness of the layer was examined at pH 5.
Abstract: The adsorption of bovine serum albumin and human serum albumin (HSA) at the air/water interface has been studied by neutron specular reflection. All the neutron measurements were performed in null reflecting water (D2O:H2O ≃ 1:11), and the specular reflectivity at this water contrast is entirely from the adsorbed protein layer at the air/water interface. Accurate measurement of surface excesses and layer thicknesses has allowed us to infer the possible structural conformation of the two protein molecules on the surface of water. The effect of bulk protein concentration on the adsorbed amount and the total thickness of the adsorbed layer was examined at pH 5, close to the isoelectric point of 4.8 for the two albumins. The surface excess (Γ) of both proteins increased sharply over the concentration range of 5 × 10-4 to 5 × 10-2 g dm-3, beyond which Γ tended to the respective saturation limits. The saturation value of surface excess of BSA at 1 g dm-3 was found to be 2.8 ± 0.3 mg m-2 as compared with 2.1 ± 0...
TL;DR: Results indicate that high-accuracy measurements are feasible for a large number of species detected simultaneously without the necessity for internal calibration and indicate the potential of such measurements, when combined with chromatographic separations, for facilitating more rapid identification of large numbers of proteins.
Abstract: The application of Fourier transform ion cyclotron resonance (FTICR) mass spectrometry to the analysis of polypeptide mixtures resulting from proteolytic digestion is described. A new 11.5-T FTICR mass spectrometer has been applied for the analysis of tryptic digestion mixtures of the protein bovine serum albumin (BSA). The improved cyclotron frequency stability and reduced frequency shifts observed over a wide range of trapped ion population sizes provide the ability to signal average spectra without degrading mass measurement accuracy, requiring internal calibration or advanced data processing schemes to compensate for variations in ion cyclotron signals brought about by different population sizes. A total of 100 spectra were signal-averaged leading to the observation of a total of 123 isotope distributions with a signal-to-noise ratio greater than 3:1. From those distributions, 86 can be ascribed to tryptic fragments of BSA on the basis of mass measurement errors of 10 ppm or less. Of these, 71 were wi...
Journal Article•
TL;DR: This study clearly demonstrated the emulsification-induced adverse events that were detrimental to the integrity of proteins and the importance of preserving protein stability toward microencapsulation.
Abstract: The objective of this study was to investigate the behavior of three proteins at an organic solvent/water interface. To simulate the first microencapsulation step of a water-in-oil-in-water emulsion technique, a water-inoil emulsion was prepared by emulsifying an aqueous protein solution in either methylene chloride or ethyl acetate. Phase separation was then followed to collect protein samples from the aqueous phase and the organic solvent/water interface. Their properties were assessed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and size exclusion-HPLC. Bovine serum albumin was relatively unharmed during emulsification, compared to other proteins such as ovalbumin and lysozyme. In particular, the methylene chloride treatment on ovalbumin led to the formation of a large quantity of water-insoluble, solid-like aggregates and changes in the composition of monomeric and dimeric ovalbumin species. With regard to the question of ovalbumin recovery, only 9.74 ∼ 37.72% of the used ovalbumin was present in the aqueous phases after emulsification. Similar penchant was noted with lysozyme. Water-insoluble aggregates brought with by emulsification were found to be covalently bound. Interestingly, less emulsification-induced denaturing effects were observed with ethyl acetate. Our study clearly demonstrated the emulsification-induced adverse events that were detrimental to the integrity of proteins and the importance of preserving protein stability toward microencapsulation.
TL;DR: Ascorbate (100 microM) almost completely prevents cigarette smoke-induced protein oxidation and thereby protects the microsomes from subsequent proteolytic degradation.
TL;DR: It is suggested that malondialdehyde could be one of the significant sources of endogenous DNA-protein crosslinks, which readily forms crosslinks between DNA and histones under physiological ionic and pH conditions.
Abstract: In an attempt to identify endogenous chemicals producing DNA-protein crosslinks, we have studied in vitro crosslinking potential of malondialdehyde, a bifunctional chemical that is ubiquitously formed as a product of lipid peroxidation of polyunsaturated fatty acids. We have found that malondialdehyde readily forms crosslinks between DNA and histones under physiological ionic and pH conditions. Formation of DNA-protein crosslinks was limited to proteins that were able to bind to DNA. Malondialdehyde failed to form DNA-protein crosslinks when histone binding to DNA was prevented by elevated ionic strength or when bovine serum albumin was used in the reaction mixture. Malondialdehyde-produced DNA-histone crosslinks were relatively stable at 37 degrees C with t1/2=13.4 days. Crosslinking of histones to DNA proceeds through the initial formation of protein adduct followed by reaction with DNA. Modification of DNA by malondialdehyde does not lead to a subsequent crosslinking of proteins. Significant formation of DNA-protein crosslinks was also registered in isolated kidney and liver nuclei treated with malondialdehyde. Based on its reactivity and stability of the resulting crosslinks, it is suggested that malondialdehyde could be one of the significant sources of endogenous DNA-protein crosslinks.
TL;DR: Upon the addition of small amounts of sodium dodecyl sulfate (SDS), the helicity of human serum albumin (HSA), lost in the urea denaturation, was mostly recovered, and the SDS denaturation finally predominates over the u Andrea denaturation.
Abstract: Upon the addition of small amounts of sodium dodecyl sulfate (SDS), the helicity of human serum albumin (HSA), lost in the urea denaturation, was mostly recovered. The profile of the recovery differed depending on the urea concentration. Then the urea concentrations were divided into three ranges: [1] a range below 3 M where the helicity only decreased as in the absence of urea (the helicity decreased down to 49% in the SDS solution); [2] a range between 4 and 8 M where the helicity initially increased up to 66% (this was the same as in the native state) and then sharply decreased; [3] a range above 9 M where the helicity only increased with an increase in added SDS concentration. When SDS was added prior to the urea denaturation, the same helicity was obtained at each surfactant concentration. Thus the SDS denaturation finally predominates over the urea denaturation. In the middle range, profiles of the structural change were rather complicated. The increase and decrease of helicity were accomplished be...