Showing papers on "Human serum albumin published in 2001"
TL;DR: Cell attachment and proliferation were surface roughness sensitive and increased as the roughness of Ti alloy increased, and may be explained by the differential adsorption of the two proteins onto smooth and rough Ti alloy surfaces.
742 citations
TL;DR: In this paper, the crystal structures at 2.5 A-resolution of HSA-myristate complexed with the R-(+) and S-(-) enantiomers of warfarin, a widely used anticoagulant, were determined.
731 citations
TL;DR: This work presents the first high-resolution crystal structures of HSA complexed with two important unsaturated fatty acids, the monounsaturated oleic acid and the polyunsaturated arachidonic acid, observed to occupy the seven binding sites distributed across the protein that are also bound by medium and long-chain saturated fatty acids.
455 citations
TL;DR: Because the degradation of the susceptible, specific transition metal binding site of HSA may account for the decreased cobalt binding observed during ischemic events, an assay that detects this reduced binding could be useful in the diagnosis of ischemia.
Abstract: Patients suffering from myocardial ischemia reportedly exhibit reduced in vitro binding of exogenous Co(2+) to the N-terminal of human serum albumin (HSA). The purpose of our investigation was to simulate changes in the N-terminus of HSA that may account for these ischemia-induced modifications to the cobalt binding site. HPLC, LC-MS and (1)H NMR analyses have shown that the N-terminal region of HSA Asp-Ala-His-Lys binds the transition metals Co(2+) and Ni(2+). Synthetic peptides with the first 2-12 amino acids of the HSA sequence demonstrated that the first three amino acids, Asp-Ala-His, are essential for strong binding of cobalt. Modification of the N-terminus peptide of HSA by way of N-acetylation or the deletion of one or more amino acid resulted in no binding of cobalt. Because the degradation of the susceptible, specific transition metal binding site of HSA may account for the decreased cobalt binding observed during ischemic events, an assay that detects this reduced binding could be useful in the diagnosis of ischemia.
285 citations
TL;DR: The overall conformation in neutral solution of BSA, as of HSA, should be rigid, in the sense indicated above, and very similar to the heart-shaped structure observed in HSA crystals.
251 citations
TL;DR: This work developed a novel, water-soluble formulation in which paclitaxel is bound noncovalently to human serum albumin, and found the Scatchard plot was found to be curvilinear with a slight positive slope of the final part.
Abstract: Paclitaxel, a very potent antitumor agent is a hydrophobic molecule with low aqueous solubility. Its currently used formula (Taxol) contains the drug in a 1 : 1 (v/v) mixture of ethanol and Cremophor EL. To minimize vehicle-related toxicity, we developed a novel, water-soluble formulation in which paclitaxel is bound noncovalently to human serum albumin. For this purpose, studies of the paclitaxel-albumin binding equilibrium were performed. Paclitaxel dissolved in ethanol was added to the aqueous solution of human serum albumin. Precipitated paclitaxel was removed and unbound drug was separated by ultrafiltration. Paclitaxel concentration was measured by RP-HPLC. Binding data were evaluated based both on the Scatchard plot and the general binding equation describing binding equilibria with the stepwise stoichiometric binding constants. The Scatchard plot was found to be curvilinear with a slight positive slope of the final part. Parameters of high affinity specific binding were determined from the initial part of the curve (nsp = 1.3 and Ksp = 1.7 x 10(6) M(-1)). Stoichiometric binding constants were estimated by fitting the general binding equation to the experimental data (K1 = 2.4 x 10(6) M(-1) and K2 = 1.0 x 10(5) M(-1)). Saturation of the protein with paclitaxel, similarly to other ligands of albumin, could not be reached. The greatest observed value of r (number of paclitaxel molecules bound to one albumin molecule) was 6.6.
198 citations
TL;DR: VOSO(4) and VO(malto)(2) showed differences when levels of plasma glucose and blood V in diabetic rodents were compared, and in the formation of V-protein complexes with abundant serum proteins, which suggest that binding of V compounds to ligands in blood, such as proteins, may affect the available pool of V for biological effects.
192 citations
TL;DR: The enantioseparations of various compounds using proteins as the chiral selectors in high-performance liquid chromatography (HPLC) are considered in this review.
192 citations
TL;DR: Mild oxidation of HSA has no detectable effect on the binding of drugs to site I in subdomain IIA and both the ligand binding property of site II and the esterase–like activity of oxidized HSAs are decreased, most probably due to conformational changes in sub domain IIIA.
Abstract: Purpose. Human serum albumin (HSA) was mildly oxidized by a metal–catalyzed oxidation system (MCO–HSA), chloramine–T (CT–HSA) or H2O2 (H2O2–HSA), and the effects of these treatments on the structural, drug–binding and esterase–like properties were studied.
Methods. Protein conformation was examined by calorimetric, chromatographic, electrophoretic and spectroscopic techniques. Drug binding was studied by ultrafiltration method, and esterase–like activity was determined using p–nitrophenyl acetate as a substrate.
Results. Far–UV and near–UV CD spectra indicated that significant structural changes had occured as the result of treatment with MCO–HSA and CT–HSA but not with H2O2–HSA. However, SDS–PAGE analysis does not provide precise information on gross conformational changes such as fragmentation, cross–linking and SDS–resistant polymerisation. The results of differential scanning calorimetry, the fluorescence of the hydrophobic probe 1,1–bis–4–anilino–naphthalene–5,5–sulfonic acid and the elution time from a hydrophobic HPLC column indicated that MCO–HSA and CT–HSA in particular, have a more open structure and a higher degree of exposure of hydrophobic areas than unoxidized HSA. In all cases, high–affinity binding of warfarin remained unchanged for all the oxidized HSAs. However, high–affinity binding of ketoprofen to CT–HSA and, especially, MCO–HSA was diminished. In addition, the esterase–like activity of these proteins were all decreased to the same low level.
Conclusions. Mild oxidation of HSA has no detectable effect on the binding of drugs to site I in subdomain IIA. In contrast, both the ligand binding property of site II and the esterase–like activity of oxidized HSAs are decreased, most probably due to conformational changes in subdomain IIIA.
133 citations
TL;DR: Present results represent a clear-cut evidence for the drug-induced shift of allosteric equilibrium(a) of HSA.
Abstract: Haem binding to human serum albumin (HSA) endows the protein with peculiar spectroscopic properties. Here, the effect of ibuprofen and warfarin on the spectroscopic properties of ferric haem-human serum albumin (ferric HSA-haem) and of ferrous nitrosylated haem-human serum albumin (ferrous HSA-haem-NO) is reported. Ferric HSA-haem is hexa-coordinated, the haem-iron atom being bonded to His105 and Tyr148. Upon drug binding to the warfarin primary site, the displacement of water molecules--buried in close proximity to the haem binding pocket--induces perturbation of the electronic absorbance properties of the chromophore without affecting the coordination number or the spin state of the haem-iron, and the quenching of the 1H-NMR relaxivity. Values of Kd for ibuprofen and warfarin binding to the warfarin primary site of ferric HSA-haem, corresponding to the ibuprofen secondary cleft, are 5.4 +/- 1.1 x 10(-4) m and 2.1 +/- 0.4 x 10(-5) m, respectively. The affinity of ibuprofen and warfarin for the warfarin primary cleft of ferric HSA-haem is lower than that reported for drug binding to haem-free HSA. Accordingly, the Kd value for haem binding to HSA increases from 1.3 +/- 0.2 x 10(-8) m in the absence of drugs to 1.5 +/- 0.2 x 10(-7) m in the presence of ibuprofen and warfarin. Ferrous HSA-haem-NO is a five-coordinated haem-iron system. Drug binding to the warfarin primary site of ferrous HSA-haem-NO induces the transition towards the six-coordinated haem-iron species, the haem-iron atom being bonded to His105. Remarkably, the ibuprofen primary cleft appears to be functionally and spectroscopically uncoupled from the haem site of HSA. Present results represent a clear-cut evidence for the drug-induced shift of allosteric equilibrium(a) of HSA.
128 citations
TL;DR: The affinity of human albumin for bilirubin is not constant, but varies with both albumin concentration and buffer composition, suggesting that binding may be considerably less avid at physiological albumin concentrations than previously believed.
TL;DR: The structure-activity relationship of the chemical series was conducted and several analogues displaying sub-nanomolar K(i) values at the EP3 receptor and micromolar activities at theEP1, EP2 and EP4 receptors were found.
TL;DR: Dynamic light scattering and turbidimetry revealed a critical pH corresponding to the onset of HA-SA soluble complex formation, and simple electrostatic considerations indicate an electrostatic binding energy for the formation of this complex of ca.
TL;DR: In this article, the solvation dynamics of 4-(dicyanomethylene)-2-methyl-6-(p-dimethylaminostyryl) 4H-pyran (DCM) in aqueous solution of a protein, human serum albumin (HSA), was studied using picosecond time-resolved emission spectroscopy.
Abstract: Solvation dynamics of 4-(dicyanomethylene)-2-methyl-6-(p-dimethylaminostyryl) 4H-pyran (DCM) in aqueous solution of a protein, human serum albumin (HSA), is studied using picosecond time-resolved emission spectroscopy. The solvation dynamics of DCM bound to HSA is found to be biexponential with one component of 600 ± 100 ps (25%) and a very long component of 10 ± 1 ns (75%). This indicates that the motion of the water molecules in the vicinity of the protein is highly constrained.
TL;DR: Two doxorubicin albumin conjugates (A-DP1 and A-DP2), which differ in their substrate specificity for the matrix metalloproteinases MMP2 and MMP9, were prepared by binding maleimide doxorbicin peptide derivatives to the cysteine-34 position of human serum albumin.
TL;DR: The results suggest a rational strategy for designing out albumin binding in potential drug molecules by using structure-based design in conjunction with NMR-based screening, and several of these compounds maintain activity against cyclooxygenase-2.
Abstract: Many lead compounds bind to serum albumin and exhibit markedly reduced efficacy in vivo as compared to their potency in vitro. To aid in the design of compounds with reduced albumin binding, we performed nuclear magnetic resonance (NMR) structural and binding studies on the complex between domain III of human serum albumin (HSA-III) and diflunisal, a cyclooxygenase inhibitor with antiinflammatory activity. The structural studies indicate that the aromatic rings of diflunisal are involved in extensive and specific interactions with hydrophobic residues that comprise the binding pocket in subdomain IIIA. The carboxylic acid of diflunisal forms electrostatic interactions with the protein similar to those observed in the X-ray structure of HSA complexed to myristic acid. In addition to the structural studies, NMR-derived binding constants were obtained for diflunisal and closely related analogues to develop a structure-affinity relationship for binding to subdomain IIIA. On the basis of the structural and binding data, compounds were synthesized that exhibit more than a 100-fold reduction in binding to domain III of HSA, and nearly a 10-fold reduction in affinity for full length albumin. Significantly, several of these compounds maintain activity against cyclooxygenase-2. These results suggest a rational strategy for designing out albumin binding in potential drug molecules by using structure-based design in conjunction with NMR-based screening.
TL;DR: The adsorption of lysozyme and human serum albumin onto hydrogel contact lenses was investigated as a function of lens surface charge and the fact that HSA was not observed on either the anionic or non-ionic charged species demonstrates the effect of charge repulsion.
TL;DR: In this article, the interaction of 7-aminocoumarins with human serum albumin (HSA) was studied by using fluorescence spectroscopic technique and modeling studies.
Journal Article•
TL;DR: In this article, the size of a human serum albumin molecule in aqueous solution containing 150 mM NaCl was studied using small-angle neutron scattering, and the molecular radius of gyration was estimated to be 27.4 +/- 0.35 A.
Abstract: The size of a human serum albumin molecule in aqueous solution containing 150 mM NaCl was studied using small-angle neutron scattering. The molecular radius of gyration was estimated to be 27.4 +/- 0.35 A. The compact sphere should have a smaller radius of gyration, whereas the popular human serum albumin model, a "cigar" 136 A long, should correspond to a greater radius of gyration. Possible shapes of the human serum albumin molecule which are in accordance with the results obtained, are the following: an extended ellipsoid less than 110 A of length or a nonsymmetrical oblate ellipsoid with a diameter of 85 A. The oblate ellipsoid might be close to the heart"-shaped structure of the crystalline human serum albumin molecule. The size of the albumin molecule does not change significantly as pH increases to 8.9. The possibility of the dynamic coexistence of various human serum albumin conformers in solution is discussed.
TL;DR: In this article, a modified polystyrene and albumin-based magnetic microspheres for red blood cell separation were modified and characterized by scanning electron and atomic force microscopy.
TL;DR: The steadystate and time-resolved fluorescence of Trp-214 was used to examine the conformation, dynamics, accessibility, thermal stability, and degree of ligand binding of human serum albumin (HSA) after entrapment of the protein in sol-gel processed glasses as mentioned in this paper.
Abstract: The steady-state and time-resolved fluorescence of Trp-214 was used to examine the conformation, dynamics, accessibility, thermal stability, and degree of ligand binding of human serum albumin (HSA) after entrapment of the protein in sol-gel processed glasses. The bioglasses were derived from tetraethyl orthosilicate and were aged in air without washing (dry-aged), in air after a washing step (washed), or in buffer (wet-aged). In all cases, significant changes were observed in the structure and dynamics of HSA, consistent with adsorption of the protein onto the silica surface combined with partial unfolding of the protein. Significant changes in the thermal stability and degree of ligand binding of the entrapped protein were also observed, with both stability and ligand binding capacity decreasing as aging continued. All proteins showed full accessibility to neutral quenchers over 2 months of aging but only partial accessibility to negatively charged quenchers, even at early aging times, indicating electrostatic repulsion of such analytes by the negatively charged matrix. Taken together, the results indicated that the reduced ligand binding for entrapped HSA was caused by a combination of protein denaturation and partial inaccessibility of the protein to negatively charged species. After 2 months of aging the entrapped proteins retained less than 15% of their binding ability in solution, regardless of which method was used to age the material. In light of these results, it is clear that improved sol-gel processing methods will be needed to overcome the time-dependent changes in the structure and function of proteins entrapped in silicate-based glasses.
TL;DR: Results of three restrained molecular dynamics simulations suggest that both lysine residues located in the IIA binding site of HSA, Lys195 and Lys199, could play a combined and comparable chemical role.
Abstract: The IIA binding site of human serum albumin (HSA) preferentially binds hydrophobic organic anions of medium size (e.g., aspirin, benzylpenicillin, warfarin, etc.) and bilirubin. This binding ability is particularly important for the distribution, metabolism, and efficacy of drugs. In addition, HSA can also covalently link to different IIA substrates owing to the presence of a highly reactive residue, Lys199, which is strategically located in the IIA site. Herein, we present results of three restrained molecular dynamics (MD) simulations of the IIA binding site on the HSA protein. From these simulations, we have determined the influence that the ionization state of the key residue, Lys199, and the nearby Lys195 has on the structure and dynamics of the IIA binding site. When Lys199 is neutral the computed average distances for the most significant interresidue contacts are in good agreement with those estimated from the X-ray coordinates. The analysis of the solvent structure and dynamics indicates that the basic form of Lys199 is likely connected to the acid form of Lys195 through a network of H-bonding water molecules with a donor --> acceptor character. The presence of these water bridges can be important for stabilizing the configuration of the IIA binding site and/or promoting a potential Lys195 --> Lys199 proton-transfer process. These results suggest that both lysine residues located in the IIA binding site of HSA, Lys195 and Lys199, could play a combined and comparable chemical role. Our simulations also give insight into the binding of bilirubin to HSA.
TL;DR: Investigation of the behavior of human serum albumin following exposure to peroxynitrite found it to be dominated by only two nitrated peptides, identified by comparison of LC retention times and collision-induced decomposition mass spectra as nitro-Y(411)TK(413) and nitro
TL;DR: P pH-profiles showed that the interaction between IS and DNSA was very sensitive to the N to B transition: “competitive-like” strong allosteric regulation was observed for binding of the two ligands to theN conformer, whereas in the B conformation binding of these molecules was nearly “independent”.
Abstract: Purpose. The study was performed for clarifying the mechanism of interaction between indoxyl sulfate (IS), a typical uremic toxin bound to site II, and site I-ligands when bound to human serum albumin (HSA). The effect of the N to B transition on the interactions was also examined.
TL;DR: In this paper, the influence of solvent systems containing poly(methoxy-polyethyleneglycol cyanoacrylate-co-nhexadecyl cyano acrylate) (PEGylated PHDCA) on the biological activity of rHuTNF-alpha was investigated.
TL;DR: This study suggests that Arg-gingipains and, to a lesser extent, Lys-gingIPain play an important role in the growth of P. gingivalis in a defined medium containing a human protein as the sole carbon and nitrogen source.
Abstract: Porphyromonas gingivalis, a bacterium associated with active chronic periodontitis lesions, produces several proteolytic enzymes that are thought to be involved in host colonization, perturbation of the immune system, and tissue destruction. The aim of the present study was to investigate the contribution of Arg- and Lys-gingipains produced by P. gingivalis to its growth. Although all of the proteins studied were degraded by P. gingivalis, only human serum albumin and transferrin supported growth during serial transfers in a chemically defined medium (CDM). Growth studies with site-directed gingipain-deficient mutants of P. gingivalis revealed that inactivation of both gingipains prevents growth, whereas inactivation of either Arg- or Lys-gingipain activity extended the doubling times to 33 or 13 h, respectively, compared to 9 h for the parent strain. Growth of the mutants and the parent strain was similar when the CDM was supplemented with a protein hydrolysate instead of human serum albumin. Incubation of resting P. gingivalis ATCC 33277 cells with fluorophore-labeled albumin indicated that the proteolytic fragments generated by the gingipains were internalized by the bacterial cells. Internalization of fluorophore-labeled albumin fragments was reduced or completely inhibited in the proteinase-deficient mutants. Interestingly, gingival crevicular fluid samples from diseased periodontal sites contained low-molecular-mass albumin fragments, whereas samples from healthy sites did not. The critical role of proteinases in the growth of P. gingivalis was further investigated using specific Arg- and Lys-gingipain inhibitors. Adding the inhibitors to CDM containing albumin revealed that leupeptin (Arg-gingipain A and B inhibitor) was more efficient at inhibiting growth than cathepsin B inhibitor II (Lys-gingipain inhibitor). Our study suggests that Arg-gingipains and, to a lesser extent, Lys-gingipain play an important role in the growth of P. gingivalis in a defined medium containing a human protein as the sole carbon and nitrogen source.
TL;DR: It is inferred that a conformation transition may occur due to the binding of the first Co(II) ion with the peptide segment of N-terminal residues 1-3, which results in a 'hinged movement' of the relatively hydrophobic 'valley' in the IA subdomain, which leads to a slow conformational transition in the albumins.
TL;DR: It showed that the matrix degradation and protein release profiles were highly polymer-dependent and the extent of burst release in the initial protein release increased with the decrease of molecular weight of PELA copolymer.
TL;DR: The findings show that Trp-214 is not as essential for the high-affinity binding of warfarin as has previously been thought.
Abstract: Correctly folded recombinant wild-type human serum albumin and the single-residue mutants K199A, W214A, R218H and H242Q were produced with the use of a yeast expression system. The changes in R218H resulted in a pronounced decrease in intrinsic fluorescence. Thermodynamic parameters for thermal denaturation of the present mutants and of five additional mutants have been determined, showing small increases in stability for two mutants (R218H and H242Q) and a larger decrease in stability for one (W214A). In the last of these, denaturation was a heterogeneous process starting at physiological temperature. The high-affinity binding constant for warfarin at pH 7.4 was determined by fluorescence spectroscopy: there was a significant increase in affinity for binding of warfarin to H242Q and K199A and a smaller decrease in affinity for W214A and R218H. The findings show that Trp-214 is not as essential for the high-affinity binding of warfarin as has previously been thought.
TL;DR: The present yeast expression system and purification procedure result in rHSA with structural and functional properties very similar to those of HSA, which showed comparable half-lives, excretion and tissue distributions of the two albumin preparations.
Abstract: Purpose. Recombinant human serum albumin (rHSA), secreted by a Pichia pastorisexpression system, was purified by a fast and efficient method, the essential feature of which is strong but reversible binding of the protein to Blue Sepharose. The structural characteristics, stability, and ligand-binding properties of the resulting protein were examined, and pre-clinical studies were performed.