Browse
Submit Data
Databases
API
Help

Dataset Information

0 Views

0 Connections

0 Citations

0 Reanalyses

0 Downloads

Omics score: 0

Multiple effects of haemin binding on protease susceptibility of bovine serum albumin and a novel isolation procedure for its large fragment.

ABSTRACT: The effects of ligand binding on the proteolytic susceptibility of BSA were investigated. The rate for proteolytic digestion with either trypsin or chymotrypsin decreased in the presence of bilirubin and fatty acids, suggesting that overall albumin conformation is stabilized by these ligands. In contrast, haemin showed multiple effects on a proteolytic digestion pattern: the rate for the degradation of intact albumin greatly increased, but a large 45 kDa fragment accumulated during proteolytic digestion in the presence of this ligand. This unique fragmentation pattern allowed us to isolate the 45 kDa fragment at a high yield (about 30% on a molar basis) by one-step purification. Sequence analyses indicated that this fragment lies between residues Thr190 and Ala583, which constitutes domains II and III of the albumin molecule. Far-u.v. c.d. spectra strongly suggested that the secondary structure in the intact albumin is almost retained in the 45 kDa fragment. The isolated 45 kDa fragment showed haemin-binding ability, as evaluated by spectroscopic titration; upon re-digestion of the 45 kDa fragment, haemin showed strong protective effects. These results were consistent with the idea that haemin binding to BSA induces an increased protease susceptibility in the loop region between domains I and II, but in the overall conformation of domains II and III, a protease-resistant property.

SUBMITTER: Shin WS

PROVIDER: S-EPMC1137455 | biostudies-other | 1994 Nov

REPOSITORIES: biostudies-other

ACCESS DATA

Json Xml

Similar Datasets

Denaturation studies on bovine serum albumin-bile salt system: Bile salt stabilizes bovine serum albumin through hydrophobicity.

Project description:Protein denaturation is under intensive research, since it leads to neurological disorders of severe consequences. Avoiding denaturation and stabilizing the proteins in their native state is of great importance, especially when proteins are used as drug molecules or vaccines. It is preferred to add pharmaceutical excipients in protein formulations to avoid denaturation and thereby stabilize them. The present study aimed at using bile salts (BSs), a group of well-known drug delivery systems, for stabilization of proteins. Bovine serum albumin (BSA) was taken as the model protein, whose association with two BSs, namely sodium cholate (NaC) and sodium deoxycholate (NaDC), was studied. Denaturation studies on the pre-formed BSA-BS systems were carried out under chemical and physical denaturation conditions. Urea was used as the chemical denaturant and BSA-BS systems were subjected to various temperature conditions to understand the thermal (physical) denaturation. With the denaturation conditions prescribed here, the data obtained is informative on the association of BSA-BS systems to be hydrophobic and this effect of hydrophobicity plays an important role in stabilizing the serum albumin in its native state under both chemical and thermal denaturation.

| S-EPMC5859114 | biostudies-literature

TALEN-mediated modification of the bovine genome for large-scale production of human serum albumin.

Project description:As an initial step towards creating genetically modified cattle as a biopharming source of recombinant human serum albumin (rHSA), we report modification of the bovine albumin (bA) locus by transcription activator-like effector nuclease (TALEN)-stimulated homology-directed repair (HDR). Pedigreed bovine fibroblasts were co-transfected with TALENs and an 11.5-kb human serum albumin (HSA) minigene donor construct, designed to simultaneously disrupt and replace bovine serum albumin (BSA) expression with controlled rHSA expression in both the liver and the milk. Targeted integration of the HSA minigene was confirmed in transfected fibroblasts at a frequency of approximately 11% and transgenic bovine embryos were produced from targeted fibroblasts using somatic cell nuclear transfer (SCNT). The research delineated here lays the foundation for the future generation of transgenic rHSA cattle with the potential to provide a large-scale, reliable, and quality-controlled source of rHSA.

| S-EPMC3931800 | biostudies-literature

Proteolysis of Iron Oxide-Associated Bovine Serum Albumin.

Project description:Proteins are a substantial nitrogen source in soils provided that they can be hydrolyzed into bioavailable small peptides or amino acids. However, the strong associations between proteins and soil minerals restrict such proteolytic reactions. This study focused on how an extracellular fungal protease (Rhizopus sp.) hydrolyzed iron oxide-associated bovine serum albumin (BSA) and the factors that affected the proteolysis. We combined batch experiments with size-exclusion and reversed phase liquid chromatography and in situ infrared spectroscopic measurements to monitor the generation of proteolytic products in solution as well as the real-time changes of the adsorbed BSA during 24 h. Results showed that protease hydrolyzed the iron oxide-associated BSA directly at the surface without an initial desorption of BSA. Concurrently, the protease was adsorbed to vacant surface sites at the iron oxides, which significantly slowed down the rate of proteolysis. This inhibiting effect was counteracted by the presence of preadsorbed phosphate or by increasing the BSA coverage, which prevented protease adsorption. Fast initial rates of iron oxide-associated BSA proteolysis, comparable to proteolysis of BSA in solution, and very slow rates at prolonged proteolysis suggest a large variability in mineral-associated proteins as a nitrogen source in soils and that only a fraction of the protein is bioavailable.

| S-EPMC7311061 | biostudies-literature

Biogenic and synthetic polyamines bind bovine serum albumin.

Project description:Biogenic polyamines are found to modulate protein synthesis at different levels, while polyamine analogues have shown major antitumor activity in multiple experimental models, including breast cancer. The aim of this study was to examine the interaction of bovine serum albumin (BSA) with biogenic polyamines, spermine and spermidine, and polyamine analogues 3,7,11,15-tetrazaheptadecane x 4 HCl (BE-333) and 3,7,11,15,19-pentazahenicosane x 5 HCl (BE-3333) in aqueous solution at physiological conditions. FTIR, UV-visible, CD, and fluorescence spectroscopic methods were used to determine the polyamine binding mode and the effects of polyamine complexation on protein stability and secondary structure. Structural analysis showed that polyamines bind BSA via both hydrophilic and hydrophobic interactions. Stronger polyamine-protein complexes formed with biogenic than synthetic polyamines with overall binding constants of K(spm) = 3.56 (+/-0.5) x 10(5) M(-1), K(spmd) = 1.77 (+/-0.4) x 10(5) M(-1), K(BE-333) = 1.11 (+/-0.3) x 10(4) M(-1) and K(BE-3333) = 3.90 (+/-0.7) x 10(4) M(-1) that correlate with their positively charged amino group contents. Major alterations of protein conformation were observed with reduction of alpha-helix from 63% (free protein) to 55-33% and increase of turn 12% (free protein) to 28-16% and random coil from 6% (free protein) to 24-17% in the polyamine-BSA complexes, indicating a partial protein unfolding. These data suggest that serum albumins might act as polyamine carrier proteins in delivering polyamine analogues to target tissues.

| S-EPMC2886673 | biostudies-literature

Kinetics and mechanism of the interaction between human serum albumin and monomeric haemin.

Project description:The interaction of human serum albumin with monomeric haemin has been investigated by detailed kinetic analysis in dimethyl sulphoxide/water (3:5, v/v). The results obtained under conditions of albumin saturation of haemin and under pseudo-single turnover conditions indicate that methaemalbumin is formed in a two-stage, single-intermediate process. The initial association between the haemin and human serum albumin is a chemically controlled process (k1 = 1.7 X 10(5) mol-1 . s-1 . dm3 at 24 degrees C); the variation of K1 with pH exhibited a well defined pK of 5.9. The overall equilibrium constant, calculated by using microscopic rate constants, is 1.1 (+/- 0.5) X 10(8) mol-1 at 24 degrees C. The data and conclusions are consistent with a general binding mechanism for albumin in which intermediate formation is followed by an entropy-controlled internalization of the ligand.

| S-EPMC1162185 | biostudies-other

Large fragments of human serum albumin.

Project description:Large fragments of human serum albumin were produced by treatment of the native protein with pepsin at pH3.5. Published sequences of human albumin [Behrens, Spiekerman & Brown (1975) Fed. Proc. Fed. Am. Soc. Exp. Biol. 34, 591; Meloun, Moravek & Kostka (1975) FEBSLett.58, 134-137]were used to locate the fragments in the primary structure. The fragments support both the sequence and proposed disulphide-linkage pattern (Behrens et al., 1975). As the pH of a solution of albumin is lowered from pH4 to pH3.5, the protein undergoes a reversible conformational change known as the N-F transition. The distribution of large fragments of human albumin digested with pepsin in the above pH region was critically dependent on pH. It appeared that this distribution was dependent on the conformation of the protein at low pH, rather than the activity of pepsin. The yields of the large fragments produced by peptic digestion at different values of pH suggested that the C-terminal region of albumin unfolds or separates from the rest of the molecule during the N-F transition. The similarity of peptic fragments of human and bovine albumin produced under identical conditions supports the proposed similar tertiary structure of these molecules.

| S-EPMC1164549 | biostudies-other

Reversible pH Responsive Bovine Serum Albumin Hydrogel Sponge Nanolayer.

Project description:A pH dependent reversible sponge like behavior of a bovine serum albumin (BSA) nanolayer adsorbed at the gold-saline interface is revealed by quartz crystal microbalance with dissipation (QCM-D), atomic force microscope (AFM) and contact angle measurements. During the saline rinsing cycles, the BSA layer adsorbs water molecules at pH 7.0 and releases them at pH 4.5. The phenomenon remains constant and reproducible upon multiple rinsing cycles. The BSA layer thickness also increases upon rinsing with saline at pH 7.0 and reverses back to its original thickness at pH 4.5. Varying ionic strength with water further desorbs more water molecules from the BSA layer, which decreases its mass and thickness. However, upon both pH and ionic strength changes, all the BSA molecules remain adsorbed irreversibly at the gold interface and only the sorption of water molecules occurs. The study aims at engineering high efficiency pH-responsive biodiagnostics and drug delivery systems.

| S-EPMC7286146 | biostudies-literature

Purification and Characterization of Bovine Serum Albumin Using Chromatographic Method.

Project description:Purpose: Albumin is an abundant protein of blood and has many biopharmaceutical applications. The aim of this study was to purify bovine serum albumin (BSA) using produced rabbit anti-BSA antibody. Methods: The polyclonal antibody was produced against the BSA in rabbits. Then, the pure BSA was injected to three white New Zealand rabbits. ELISA test was done to evaluate antibody production. After antibody purification,the purified antibody was attached to CNBr-activated sepharose and finally it was used for purification of albumin from bovine serum. Western blotting analysis was used for functional assessment of immunoaffinity purified BSA. Results: The titer of anti-bovine albumin determined by ELISA was obtained 1: 256000. The SDS-PAGE showed up to 98% purity of isolated BSA and western blotting confirmed the BSA functionality. Purified bovine serum albumin by affinity chromatography showed a single band with molecular weight of 66 KDa. Conclusion: Affinity chromatography using produced rabbit anti-BSA antibody would be an economical and safe method for purification of BSA.

| S-EPMC5241424 | biostudies-literature

Binding Constants of Substituted Benzoic Acids with Bovine Serum Albumin.

Project description:Experimental data on the affinity of various substances to albumin are essential for the development of empirical models to predict plasma binding of drug candidates. Binding of 24 substituted benzoic acid anions to bovine serum albumin was studied using spectrofluorimetric titration. The equilibrium constants of binding at 298 K were determined according to 1:1 complex formation model. The relationships between the ligand structure and albumin affinity are analyzed. The binding constant values for m- and p-monosubstituted acids show a good correlation with the Hammett constants of substituents. Two- and three-parameter quantitative structure-activity relationship (QSAR) models with theoretical molecular descriptors are able to satisfactorily describe the obtained values for the whole set of acids. It is shown that the electron-density distribution in the aromatic ring exerts crucial influence on the albumin affinity.

| S-EPMC7169394 | biostudies-literature

Kinetics of Thermal Denaturation and Aggregation of Bovine Serum Albumin.

Project description:Thermal aggregation of bovine serum albumin (BSA) has been studied using dynamic light scattering, asymmetric flow field-flow fractionation and analytical ultracentrifugation. The studies were carried out at fixed temperatures (60°C, 65°C, 70°C and 80°C) in 0.1 M phosphate buffer, pH 7.0, at BSA concentration of 1 mg/ml. Thermal denaturation of the protein was studied by differential scanning calorimetry. Analysis of the experimental data shows that at 65°C the stage of protein unfolding and individual stages of protein aggregation are markedly separated in time. This circumstance allowed us to propose the following mechanism of thermal aggregation of BSA. Protein unfolding results in the formation of two forms of the non-native protein with different propensity to aggregation. One of the forms (highly reactive unfolded form, Uhr) is characterized by a high rate of aggregation. Aggregation of Uhr leads to the formation of primary aggregates with the hydrodynamic radius (Rh,1) of 10.3 nm. The second form (low reactive unfolded form, Ulr) participates in the aggregation process by its attachment to the primary aggregates produced by the Uhr form and possesses ability for self-aggregation with formation of stable small-sized aggregates (Ast). At complete exhaustion of Ulr, secondary aggregates with the hydrodynamic radius (Rh,2) of 12.8 nm are formed. At 60°C the rates of unfolding and aggregation are commensurate, at 70°C the rates of formation of the primary and secondary aggregates are commensurate, at 80°C the registration of the initial stages of aggregation is complicated by formation of large-sized aggregates.

| S-EPMC4839713 | biostudies-literature

OmicsDI is part of the ELIXIR infrastructure

OmicsDI is an Elixir interoperability service. Learn more ›

Tweets

OmicsDI Databases

PRIDE
PeptideAtlas
MassIVE
JPOST Repository
Physiome Model Repository

EGA
EVA
ENA
LINCS
PAXDB
Cell Collective

MetaboLights
Metabolomics Workbench
MetabolomeExpress
GNPS
BioModels
FAIRDOMHub

ArrayExpress
dbGaP
ExpressionAtlas
GEO
NODE

Information

Databases
Help
API
Contact us
Code on GitHub
Terms of use
Submit Data