Showing papers on "Human serum albumin published in 2009"
TL;DR: A sensor based on a hybrid synthetic-biomolecule that uses arrays of green fluorescent protein and nanoparticles to detect proteins at biorelevant concentrations in both buffer and human serum is reported.
Abstract: There is a direct correlation between protein levels and disease states in human serum, which makes it an attractive target for sensors and diagnostics. However, this is challenging because serum features more than 20,000 proteins, with an overall protein content greater than 1 mM. Here we report a sensor based on a hybrid synthetic-biomolecule that uses arrays of green fluorescent protein and nanoparticles to detect proteins at biorelevant concentrations in both buffer and human serum. Distinct and reproducible fluorescence-response patterns were obtained from five serum proteins (human serum albumin, immunoglobulin G, transferrin, fibrinogen and a-antitrypsin), both in buffer and when spiked into human serum. Using linear discriminant analysis we identified these proteins with an identification accuracy of 100% in buffer and 97% in human serum. The arrays were also able to discriminate between different concentrations of the same protein, as well as a mixture of different proteins in human serum.
425 citations
TL;DR: It was proved that the fluorescence quenching of HSA by berberine is a result of the formation of berberin-HSA complex and the results of thermodynamic parameters DeltaG, DeltaH, and DeltaS at different temperatures indicate that the electrostatic interactions play a major role for berberines- HSA association.
404 citations
01 Jan 2009
TL;DR: A review on the binding strategies of protein to various ligands that may be expected to lead to significant clinical applications is presented in this paper, where the authors highlight current overview of protein binding strategies.
Abstract: Human serum albumin (HSA), being the most abundant carrier protein in blood and a modern day clinical tool for drug delivery, attracts high attention among biologists. Hence, its unfolding/refolding strategies and exogenous/endogenous ligand binding preference are of immense use in therapeutics and clinical biochemistry. Among its fellow proteins albumin is known to carry almost every small molecule. Thus, it is a potential contender for being a molecular cargo/or nanovehicle for clinical, biophysical and industrial purposes. Nonetheless, its structure and function are largely regulated by various chemical and physical factors to accommodate HSA to its func- tional purpose. This multifunctional protein also possesses enzymatic properties which may be used to convert prodrugs to active therapeutics. This review aims to highlight current overview on the binding strategies of protein to various ligands that may be expected to lead to significant clinical applications. Chirality 00:000-000, 2009. V C 2009 Wiley-Liss, Inc.
284 citations
TL;DR: Since further progress is likely to benefit from increased structural scrutiny of HSA, methodological developments instrumental to the success of crystallographic analysis of the protein are discussed in some detail.
255 citations
TL;DR: Structural analysis showed that HSA conformation was altered by dendrimers with a major reduction of alpha-helix and increase in random coil and turn structures suggesting a partial protein unfolding.
Abstract: Dendrimers are synthetic, highly branched, spherical macromolecules with nanometer dimensions and potential applications in DNA and drug delivery systems. Human serum albumin (HSA) is a major transporter for delivering several endogenous compounds and drugs in vivo. The aim of this study was to examine the interaction of human serum albumin with several dendrimers such as mPEG-PAMAM (G3), mPEG-PAMAM (G4), and PAMAM (G4) at physiological conditions, using constant protein concentration and various dendrimer compositions. FTIR, UV-visible, CD, and fluorescence spectroscopic methods were used to analyze macromolecule binding mode, the binding constant and the effects of dendrimers complexation on HSA stability and conformation. Structural analysis showed that dendrimers bind HSA via polypeptide polar groups (hydrophilic) with number of bound polymer (n) 1.08 (mPEG-PAMAM-G3), 1.50 (mPEG-PAMAM-G4), and 0.96 (PAMAM-G4). The overall binding constants estimated were of KmPEG-G3=1.3 (+/-0.2)x10(4) M(-1), KmPEG-G4=2.2 (+/-0.4)x10(4) M(-1), and KPAMAM-G4=2.6 (+/-0.5)x10(4) M(-1). HSA conformation was altered by dendrimers with a major reduction of alpha-helix and increase in random coil and turn structures suggesting a partial protein unfolding.
205 citations
TL;DR: The results of this study suggest marked dysfunction of albumin function in advanced cirrhosis and provide further evidence for damage to the circulating albumin, which is not reversed by MARS therapy.
200 citations
TL;DR: Structural analysis showed that curcumin and genistein bind HSA via polypeptide polar groups with overall binding constants of K(curcumin) and K(genistein) and the HSA conformation was altered by pigment complexation with reduction of alpha-helix and increase of random coil and turn structures suggesting a partial protein unfolding.
188 citations
TL;DR: The results support that strong binding occurs at the hydrophobic moieties of HSA and fibrinogen, excluding water access, and lead to effective suppression of degradation of curcumin.
Abstract: The use of curcumin as an effective wound healing agent is of significant interest currently. It is well established that curcumin undergoes rapid degradation in physiological buffer by hydrolysis. The means by which curcumin is stabilized at the wound site to enable healing is poorly understood because blood plasma is composed of approximately 92% water. Plasma proteins, which constitute the remaining 6-8%, has been shown to stabilize curcumin. It is, however, still unclear which proteins are responsible for this phenomenon. In this study, the effects of major plasma proteins, which include human serum albumin (HSA), fibrinogen, immunoglobulin G (IgG), and transferrin, on stabilizing curcumin are investigated. In particular, we investigate their effects on the hydrolysis of curcumin at pH 7.4. In the presence of both transferrin and IgG, curcumin continues to undergo rapid hydrolysis but this reaction is suppressed by the presence of either HSA or fibrinogen with an impressive yield of approximately 95%. Furthermore, the binding constants of curcumin to HSA and fibrinogen are on the order of 10(4) and 10(5) M(-1), respectively. The binding constants of transferrin and IgG, however, are at least 1 order of magnitude less than those of HSA and fibrinogen. The results support that strong binding occurs at the hydrophobic moieties of HSA and fibrinogen, excluding water access. Therefore, strong interactions with HSA and fibrinogen inhibit hydrolysis of curcumin and in turn lead to effective suppression of degradation.
179 citations
TL;DR: HSA conformation was stabilized by cholesterol and DOPE with a slight increase of protein alpha-helical structures, while DOTAP and DDAB induced an important (alpha-->beta) transition, suggesting a partial protein unfolding.
Abstract: Human serum albumin (HSA) is a major transporter for delivering several endogenous compounds including fatty acids in vivo. Even though HSA is the primary target of fatty acid binding, the effects of cationic lipid on protein stability and conformation have not been investigated. The aim of this study was to examine the interaction of human serum albumin (HSA) with helper lipids--cholesterol (Chol) and dioleoylphosphatidylethanolamine (DOPE)--and with cationic lipids--dioctadecyldimethylammonium bromide (DDAB) and 1,2-dioleoyl-3-trimethylammonium-propane (DOTAP), at physiological conditions, using constant protein concentration and various lipid contents. Fourier transform infrared (FTIR), circular dichroism (CD), and fluorescence spectroscopic methods were used to analyze the lipid binding mode, the binding constant, and the effects of lipid interaction on HSA stability and conformation. Structural analysis showed that cholesterol and DOPE (helper lipids) interact mainly with HSA polypeptide polar groups and via hydrophobic moieties. Hydrophobic interactions dominate the binding of cationic lipids to HSA. The number of bound lipids (n) calculated was 1.22 (cholesterol), 1.82 (DDAB), 1.76 (DOPE), and 1.56 (DOTAP). The overall binding constants estimated were KChol=2.3 (+/-0.50)x10(3) M(-1), KDDAB=8.9 (+/-0.95)x10(3) M(-1), KDOTAP=9.1 (+/-0.90)x10(3) M(-1), and KDOPE=4.7 (+/-0.70)x10(3) M(-1). HSA conformation was stabilized by cholesterol and DOPE with a slight increase of protein alpha-helical structures, while DOTAP and DDAB induced an important (alpha-->beta) transition, suggesting a partial protein unfolding.
177 citations
TL;DR: The movement of intravitreal nanoparticles depended on both nanoparticles surface charge and retinal injury, and the anionic HSA-NP is a promising drug or gene delivery carrier to the sub-retinal space and RPE.
Abstract: Purpose
To investigate the movement of intravitreally injected human serum albumin nanoparticles (HSA-NP) with respect to nanoparticle surface charge and retinal injury.
162 citations
TL;DR: Binding of five perfluoroalkyl acids with human serum albumin (HSA) was investigated by site-specific fluorescence and the data were compared with fatty acids to evaluate the potential toxicological effect of these environmental chemicals.
Abstract: Binding of five perfluoroalkyl acids with human serum albumin (HSA) was investigated by site-specific fluorescence. Intrinsic fluorescence of tryptophan-214 in HSA was monitored upon addition of the chemicals. Although perfluorobutyl acid (PFBA) and perfluorobutane sulfonate (PFBS) did not cause fluorescence change, perfluorooctanoic acid (PFOA), perfluorooctane sulfonate (PFOS), and perfluorododecanoic acid (PFDoA) induced fluorescence quenching, from which binding constant of 2.7 x 10(5) M(-1) for PFOA and 2.2 x 10(4) M(-1) for PFOS was calculated. Two fluorescent probes, dansylamide (DA) and dansyl-L: -proline (DP), were employed in fluorescence displacement measurements to study the interaction at two Sudlow's binding sites. At Site I, both PFBA and PFBS displaced DA with binding constants of 1.0 x 10(6) M(-1) and 2.2 x 10(6) M(-1). At Site II, PFBS and PFDoA displaced DP with binding constants of 6.5 x 10(6) M(-1) and 1.2 x 10(6) M(-1), whereas PFBA did not bind. The data were compared with fatty acids to evaluate the potential toxicological effect of these environmental chemicals.
TL;DR: The experimental results showed that the fluorescence quenching of HSA by IMI was a result of the formation of IMI-HSA complex; staticQuenching was confirmed to result in the fluorescent quenched.
Abstract: The interaction between imidacloprid (IMI) and human serum albumin (HSA) was investigated using fluorescence and UV/vis absorption spectroscopy. The experimental results showed that the fluorescence quenching of HSA by IMI was a result of the formation of IMI-HSA complex; static quenching was confirmed to result in the fluorescence quenching. The apparent binding constant K(A) between IMI and HSA at three differences were obtained to be 1.51 x 10(4), 1.58 x 10(4), and 2.19 x 10(4) L mol(-1), respectively. The thermodynamic parameters, DeltaH degrees and DeltaS degrees were estimated to be 28.44 kJ mol(-1), 174.76 J mol(-1) K(-1) according to the van't Hoff equation. Hydrophobic interactions played a major role in stabilizing the complex. The distance r between donor (HSA) and acceptor (IMI) was obtained according to fluorescence resonance energy transfer. The effect of IMI on the conformation of HSA was analyzed using synchronous fluorescence spectroscopy CD and three-dimensional fluorescence spectra, the environment around Trp and Tyr residues were altered.
TL;DR: The finding that promiscuous protein scaffolds become sinks for holes and electrons when holding light-activated pigments provides a new insight to the evolution and action mechanism of natural light activated oxidoreductases and new guidelines for the preparation of synthetic-light converting machineries.
Abstract: Light-induced radical generation is the hallmark of fundamental processes and many applications including photosynthesis and photodynamic therapy (PDT). In this manuscript, we present two novel observations made upon monitoring light-induced generation of reactive oxygen species (ROS) in aqueous solutions by WST11, a water-soluble derivative of the photosynthetic pigment Bacteriochlorophyll a (Bchl). Using a host of complementary experimental techniques including time-resolved spectroscopy at the subpicosecond to the millisecond range, ESR spectroscopy, electrochemistry, spectroelectrochemistry, oximetry, and protein mass spectroscopy, we first show that in aqueous solutions WST11 generates only superoxide (O(2)(-*)) and hydroxyl (OH*) radicals with no detectable traces of singlet oxygen. Second, we show that WST11 makes a noncovalent complex with human serum albumin (HSA) and that this complex functions as a photocatalytic oxidoreductase at biologically relevant concentrations enabling approximately 15 cycles of electron transfer from the associated HSA protein to molecular oxygen in the solution. These findings rule out the paradigm that porphyrin and chlorophyll based PDT is mainly mediated by formation of singlet oxygen, particularly in vascular targeted photodynamic therapy (VTP) with sensitizers that undergo photoactivation during circulation in the plasma, like [Pd]-Bacteriopheophorbide (WST09, Tookad). At the same time, our findings open the way for new design paradigms of novel sensitizers, since O(2)(-*) and OH* radicals are well-recognized precursors of important pathophysiological processes that can be activated for achieving tumor eradication. Moreover, the finding that promiscuous protein scaffolds become sinks for holes and electrons when holding light-activated pigments provides a new insight to the evolution and action mechanism of natural light activated oxidoreductases (such as photosynthetic reaction centers) and new guidelines for the preparation of synthetic-light converting machineries.
TL;DR: The interaction of a potent synthetic derivative of curcumin, isoxazolcurcumin (IOC) with human serum albumin (HSA) is reported using various biophysical methods and reveals that the interaction is entropy driven with predominantly hydrophobic forces.
Abstract: Curcumin is a nontoxic natural product with diverse pharmacological potencies. We report the interaction of a potent synthetic derivative of curcumin, isoxazolcurcumin (IOC) with human serum albumin (HSA) using various biophysical methods. The observed fluorescence quenching of HSA by IOC is due to a complex formation by a static quenching process with a quenching constant of the order of 10(5) M(-1). The binding affinity and the number of binding sites were obtained from a Scatchard analysis. Thermodynamics reveals that the interaction is entropy driven with predominantly hydrophobic forces. From the observed Forster-type fluorescence resonance energy transfer (FRET), the donor (Trp 214 in HSA) to acceptor (IOC) distance is calculated to be 3.2 nm. The conformational changes of HSA due to the interaction were investigated qualitatively from synchronous fluorescence spectra along with a quantitative estimation of the secondary structure from Fourier Transform Infrared (FTIR) and circular dichroism (CD) spectroscopies. Molecular docking studies were performed to obtain information on the possible residues involved in the interaction process, and changes in accessible surface area of the interacting residues were calculated. The preferred binding site of IOC was analyzed by ligand displacement experiments with 1-anilino-8-naphthalenesulfonate (ANS) and warfarin-bound HSA.
TL;DR: The receptor specific interactions of GNR-BBN conjugates provide realistic opportunities in the design and development of in vivo molecular imaging and therapy agents for cancer.
Abstract: Gastrin releasing protein receptor specific bombesin (BBN) peptide-gold nanoconjugates were successfully synthesized using gold nanorods and dithiolated peptide. The gold nanorod-bombesin (GNR-BBN) conjugates showed extraordinary in vitro stabilities against various biomolecules including NaCl, cysteine, histidine, bovine serum albumin, human serum albumin, and dithiothreitol. Quantitative measurements on the binding affinity (IC(50)) of GNR-BBN conjugates toward prostate and breast tumor cells were evaluated. The IC(50) values establish that GNR-BBN conjugates have strong affinity toward the gastrin releasing peptide receptors on both the tumors. Detailed cellular interaction studies of GNR-BBN conjugates revealed that nanorods internalize via a receptor-mediated endocytosis pathway. The receptor specific interactions of GNR-BBN conjugates provide realistic opportunities in the design and development of in vivo molecular imaging and therapy agents for cancer.
TL;DR: Interestingly, human serum albumin dimerizes in response to [bmim]Cl, whereas cytochrome c remains predominantly in monomeric form, which suggests a facile pathway through which biocatalytic activity can be altered in these nascent and potentially green electrolyte systems.
TL;DR: It is suggested that micro TOF-Q mass spectrometry is useful technique for drug binding studies after showing for the first time that BA isolated from a plant (Tephrosia calophylla) binds to HSA.
Abstract: Betulinic acid (BA) has anti cancer and anti-HIV activity and has been proved to be therapeutically effective against cancerous and HIV-infected cells. Human serum albumin (HSA) is the predominant protein in the blood. Most drugs that bind to HSA will be transported to other parts of the body. Using micro TOF-Q mass spectrometry, we have shown, for the first time that BA isolated from a plant (Tephrosia calophylla) binds to HSA. The binding constant of BA to HSA was calculated from fluorescence data and found to be KBA = 1.685 ± 0.01 × 106 M−1, indicating a strong binding affinity. The secondary structure of the HSA–BA complex was determined by circular dichroism. The results indicate that the HSA in this complex is partially unfolded. Further, binding of BA at nanomolar concentrations of BA to free HSA was detected using micro TOF-Q mass spectrometry. The study revealed a mass increase from 65199 Da (free HSA) to 65643 Da (HSA + drug), where the additional mass of 444 Da was due to bound BA. Based on the results of this study, it is suggested that micro TOF-Q mass spectrometry is useful technique for drug binding studies.
TL;DR: The results show that for immunoglobulins a direct and strong polysorbate binding to the protein is not the reason for the colloidal stabilisation effect of immunoglobeulins in solution in the presence of poly sorbate 20 or 80, thus showing that the thermal protein stability is quite low and negligible.
TL;DR: It is observed that fibril formation is largely affected by electrostatic shielding: at physiological pH, fibrilation is progressively more efficient and faster in the presence of up to 50 mM NaCl; meanwhile, at larger salt concentrations, excessive shielding and further enhancement of the solution hydrophobicity might involve a change in the energy landscape of the aggregation process, which makes the fibrillation process difficult.
Abstract: The fibrillation propensity of the multidomain protein human serum albumin (HSA) has been analyzed under physiological and acidic conditions at room and elevated temperatures with varying ionic strengths by different spectroscopic techniques. The kinetics of fibril formation under the different solution conditions and the structures of resulting fibrillar aggregates were also determined. In this way, we have observed that fibril formation is largely affected by electrostatic shielding: at physiological pH, fibrillation is progressively more efficient and faster in the presence of up to 50 mM NaCl; meanwhile, at larger salt concentrations, excessive shielding and further enhancement of the solution hydrophobicity might involve a change in the energy landscape of the aggregation process, which makes the fibrillation process difficult. In contrast, under acidic conditions, a continuous progressive enhancement of HSA fibrillation is observed as the electrolyte concentration in solution increases. Both the distinct ionization and initial structural states of the protein before incubation may be the origin of this behavior. CD, FT-IR, and tryptophan fluorescence spectra seem to confirm this picture by monitoring the structural changes in both protein tertiary and secondary structures along the fibrillation process. On the other hand, the fibrillation of HSA does not show a lag phase except at pH 3.0 in the absence of added salt. Finally, differences in the structure of the intermediates and resulting fibrils under the different conditions are also elucidated by TEM and FT-IR.
Patent•
10 Apr 2009
TL;DR: In this article, a human serum albumin (HSA) linker and HSA linker with binding, diagnostic, and therapeutic agents conjugated thereto is presented, which is useful in reducing tumor cell proliferation, e.g., for therapeutic therapeutic applications.
Abstract: Disclosed is a human serum albumin (HSA) linker and HSA linker with binding, diagnostic, and therapeutic agents conjugated thereto. Also disclosed is a conjugate in which the HSA linker is covalently bonded to amino and carboxy terminal binding moieties that are first and second single-chain Fv molecules (scFvs). Exemplified conjugates are useful, e.g., in reducing tumor cell proliferation, e.g., for therapeutic therapeutic applications. Also disclosed are methods and kits for the diagnostic and therapeutic application of an HSA linker conjugate.
TL;DR: The visible CD spectra indicated that the optical activity observed during the ligand binding due to induced-protein chirality suggested the important role of the phenolic OH group of CUR in the binding process.
Abstract: The current study reports the binding of curcumin (CUR) as the main pharmacologically active ingredient of turmeric and diacetylcurcumin (DAC) as a bioactive derivative of curcumin to human serum albumin (HSA) and bovine serum albumin (BSA). The apparent binding constants and number of substantive binding sites have been evaluated by fluorescence quenching method. The distance (r) between donor (HSA and BSA) and acceptor (CUR and DAC) was obtained on the basis of the Forster’s theory of non-radiative energy transfer. The minor changes on the far-UV circular dichroism spectra resulted in partial changes in the calculated secondary structure contents of HSA and BSA. The negligible alteration in the secondary structure of both albumin proteins indicated that ligand-induced conformational changes are localized to the binding site and do not involve considerable changes in protein folding. The visible CD spectra indicated that the optical activity observed during the ligand binding due to induced-protein chirality. All of the achieved results suggested the important role of the phenolic OH group of CUR in the binding process.
TL;DR: In this article, the interactions of various insulin mimetic oxovanadium(IV) compounds with serum proteins were studied in model systems and in ex vivo samples. And the results were confirmed by HPLC-ICP-MS (liquid chromatography-inductively coupled plasma spectroscopy mass spectrometry) measurements.
Abstract: The interactions of various insulin mimetic oxovanadium(IV) compounds with serum proteins were studied in model systems and in ex vivo samples. For the modeling study, an earlier in situ method was extended and applied to the formation of ternary complexes of apotransferrin (apoTf)–VIVO–maltol (mal) and 1,2-dimethyl-3-hydroxy-4(1H)-pyridinone (dhp). Both systems were evaluated via simultaneous CD and EPR measurements. Determination of the formation constants of the ternary complexes allowed the calculation of more accurate stability constants for the VIVO–apoTf parent complexes and establishment of a better model for drug speciation in serum. It was found that dhp and the synergistic carbonate are non-competitive binders. Based on the stability constants obtained for VIVO–apoTf complexes and estimated for VIVO–HSA (= human serum albumin), modeling calculations were performed on the distribution of VIVO among the components of blood serum. The results were confirmed by HPLC-ICP-MS (liquid chromatography-inductively coupled plasma spectroscopy-mass spectrometry) measurements. The ex vivo interactions of the VIVO complexes formed with mal, picolinic acid (pic) and dhp with serum protein standards and also with human serum samples were evaluated. The proteins were firstly separated by (HPLC), and the V content of each fraction was determined by ICP-MS. All the studied VIVO compounds displayed similar chromatographic profiles, associated almost exclusively with apotransferrin as predicted by the modeling calculations. Under physiological conditions, the interactions with HSA of all of the species under study were negligible. Therefore Tf seems to be the main VIVO transporter in the serum under in vitro conditions, and this association is practically independent of the chemical form in which VIVO is administered.
TL;DR: High-exclusion high-performance liquid chromatographic analysis of human serum albumin showed at least two peaks, the principal component corresponding to human mercaptalbumin (HMA) and the second one to human nonmercaptalbumin(HNA), which might be due to weak resin-HSA interaction.
Abstract: Gel-exclusion high-performance liquid chromatographic (HPLC) analysis of human serum albumin (HSA) on PGP 2000 column (0.10 M sodium phosphate buffer, 0.30 M NaCl, pH 6.86) showed at least two peaks, the principal component corresponding to human mercaptalbumin (HMA) and the second one to human nonmercaptalbumin (HNA). Mechanism for the separation of HMA and HNA might be due to weak resin-HSA interaction. HPLC analysis of bovine plasma albumin (BPA) showed a single peak on PGP 2000 column. The elution volume of HSA was larger than that of BPA, resulting in a clear resolution of HSA and BPA.
TL;DR: Spectroscopic analysis of the emission quenching at different temperatures has revealed that the quench mechanism of oxaliplatin with HSA was static quenched mechanism.
TL;DR: By the analysis of fluorescence intensity, it was observed that Pd(II) complexes have strong abilities to quench the intrinsic fluorescence of HSA through a dynamic quenching procedure and indicated that hydrophobic forces play a major role in the interaction of Bipyridin-Pd( II) complexes with HSA.
Abstract: The pharmacokinetics and pharmacodynamics of any drug will depend, largely, on the interaction that it has with human serum albumin (HSA), the most abundant plasma protein. The interaction between newly synthesized Pd(II) complexes, 2,2′-bipyridin octyl dithiocarbamato Pd(II) nitrate (Octpd), 2,2′-bipyridin butyl dithiocarbamato Pd(II) nitrate (ButPd), 2,2′-bipyridin ethyl dithiocarbamato Pd(II) nitrate (EtPd), antitumor components, with human serum albumin, a carrier protein, were studied at different temperatures of 27 and 37 °C by fluorescence spectroscopy, far UV circular dichroism (CD), and spectrophotometric and differential scanning calorimetry (DSC) techniques. By the analysis of fluorescence intensity, it was observed that Pd(II) complexes have strong abilities to quench the intrinsic fluorescence of HSA through a dynamic quenching procedure. The binding parameters were evaluated by the fluorescence quenching method. The thermodynamic parameters, including ΔH°, ΔS°, and ΔG°, were calculated by th...
TL;DR: It is concluded that the most likely fate for sulfenic acid formed in the plasma environment is the reaction with low molecular weight thiols to form mixed disulfides, a reversible modification that is actually observed in ~25% of circulating albumin.
Abstract: Human serum albumin (HSA) is the most abundant protein in the intravascular compartment. It possesses a single thiol, Cys34, which constitutes ~80% of the total thiols in plasma. This thiol is able to scavenge plasma oxidants. A central intermediate in this potential antioxidant activity of human serum albumin is sulfenic acid (HSA-SOH). Work from our laboratories has demonstrated the formation of a relatively stable sulfenic acid in albumin through complementary spectrophotometric and mass spectrometric approaches. Recently, we have been able to obtain quantitative data that allowed us to measure the rate constants of sulfenic acid reactions with molecules of analytical and biological interest. Kinetic considerations led us to conclude that the most likely fate for sulfenic acid formed in the plasma environment is the reaction with low molecular weight thiols to form mixed disulfides, a reversible modification that is actually observed in ~25% of circulating albumin. Another possible fate for sulfenic acid is further oxidation to sulfinic and sulfonic acids. These irreversible modifications are also detected in the circulation. Oxidized forms of albumin are increased in different pathophysiological conditions and sulfenic acid lies in a mechanistic junction, relating oxidizing species to final thiol oxidation products.
TL;DR: Three distinct Co(II) binding sites in HSA are characterized using circular dichroism spectroscopic titrations to confirm the presence of these three binding sites and determine their conditional binding constants at pH 7.4.
TL;DR: Investigation of the precise binding region of this bacterial receptor by protein engineering applying an alanine‐scanning procedure followed by binding studies by surface plasmon resonance (SPR) shows that the HSA binding involves residues mainly in the second α‐helix.
Abstract: Streptococcal protein G (SpG) is a bacterial cell surface receptor exhibiting affinity to both human immunoglobulin (IgG) and human serum albumin (HSA). Interestingly, the serum albumin and immunoglobulin-binding activities have been shown to reside at functionally and structurally separated receptor domains. The binding domain of the HSA-binding part has been shown to be a 46-residue triple α-helical structure, but the binding site to HSA has not yet been determined. Here, we have investigated the precise binding region of this bacterial receptor by protein engineering applying an alanine-scanning procedure followed by binding studies by surface plasmon resonance (SPR). The secondary structure as well as the HSA binding of the resulting albumin-binding domain (ABD) variants were analyzed using circular dichroism (CD) and affinity blotting. The analysis shows that the HSA binding involves residues mainly in the second α-helix.
TL;DR: The HSA/Abeta oligomer complexes have K(D) values in the micromolar to submicromolar range and compete with the further addition of Abeta monomers to the Abeta assemblies, thus inhibiting fibril growth ("monomer competitor" model).
TL;DR: There is a close relationship between oxidative stress and carbonyl stress and these are enhanced in correlation with the level of renal dysfunction among predialysis CKD patients.
Abstract: Oxidative stress is enhanced in advanced chronic kidney disease (CKD) patients and recognized as a main contributor to cardiovascular disease. Carbonyl stress is also known to be enhanced in advanced CKD; however the precise relationship between oxidative stress and carbonyl stress is not clear. The aim of this study was to investigate potential relationships between oxidative stress, carbonyl stress, and renal function among predialysis patients with CKD. A total of 32 predialysis CKD patients (22 male, 10 female) were divided into four groups according to their values for creatinine clearance (Ccr) (group A, ≥60 ml/min; group B, 45–59 ml/min; group C, 30–44 ml/min; group D, ≤29 ml/min). As main markers of oxidative and carbonyl stresses, the redox state of Cys-34 (free thiol group) of human serum albumin [HSA(Cys-34)-redox] and the carbonyl content of serum proteins were employed, respectively. The values for the fraction of both reversibly oxidized HSA [f(HNA-1)] and irreversibly oxidized HSA [f(HNA-2)] significantly increased with a decrease in renal function (group A, 21.0 ± 3.4 and 1.8 ± 0.3%; group D, 31.1 ± 4.1 and 2.7 ± 0.9%, respectively). The value for carbonyl content also significantly increased with a decrease in renal function (group A, 0.7 ± 0.1 nmol/mg protein; group D, 1.1 ± 0.2 nmol/mg protein). There was a significant positive correlation between carbonyl content and the f(HNA-2) value, while such a correlation was not observed between carbonyl content and the f(HNA-1) value, suggesting that there is a close relationship between serum protein carbonylation and irreversible albumin thiol oxidation. There is a close relationship between oxidative stress and carbonyl stress and these are enhanced in correlation with the level of renal dysfunction among predialysis CKD patients.