Changes Of Human Serum Albumin Research Articles

Albumin Oxidation and Albumin Glycation Discordance During Type 2 Diabetes Therapy: Biological and Clinical Implications.

Aims: Cys34 albumin redox modifications (reversible "cysteinylation" and irreversible "di/trioxidation"), besides being just oxidative stress biomarkers, may have primary pathogenetic roles to initiate and/or aggravate cell, tissue, and vascular damage in diabetes. In an exploratory "proof-of-concept" pilot study, we examined longitudinal changes in albumin oxidation during diabetes therapy. Methods: Mass spectrometric analysis was utilized to monitor changes in human serum albumin (HSA) post-translational modifications {glycation [glycated albumin (GA)], cysteinylation [cysteinylated albumin (CA) or human non-mercaptalbumin-1; reversible], di/trioxidation (di/trioxidized albumin or human non-mercaptalbumin-2; irreversible), and truncation (truncated albumin)} during ongoing therapy. Four informative groups of subjects were evaluated [type 1 diabetes (T1DM), type 2 diabetes (T2DM), prediabetes-obesity, and healthy controls] at baseline, and subjects with diabetes were followed for a period up to 280 days. Results: At baseline, T2DM was associated with relatively enhanced albumin cysteinylation (CA% total) compared with T1DM (P = 0.004), despite comparable mean hyperglycemia (P values: hemoglobin A1c = 0.09; GA = 0.09). T2DM, compared with T1DM, exhibited selectively and significantly higher elevations of all the "individual" glycated cum cysteinylated ("multimodified") albumin isoforms (P values: CysHSA+1G = 0.003; CysHSA+2G = 0.007; and CysHSA+3G = 0.001). Improvements in glycemic control and decreases in albumin glycation during diabetes therapy in T2DM were not always associated with concurrent reductions of albumin cysteinylation, and in some therapeutic situations, albumin cysteinylation worsened (glycation-cysteinylation discordance). Important differences were observed between the effects of sulfonylureas and metformin on albumin molecular modifications. Conclusions: T2DM was associated with higher oxidative (cysteinylation) and combined (cysteinylation plus glycation) albumin molecular modifications, which are not ameliorated by improved glucose control alone. Further studies are required to establish the clinical significance and optimal therapeutic strategies to address oxidative protein damage and resulting consequences in diabetes.

Structure of Human Serum Albumin at a Foam Surface.

Proteins can be adsorbed on the air-water interface (AWI), and the structural changes in proteins at the AWI are closely related to the foaming properties of foods and beverages. However, how these structural changes in proteins at the AWI occur is not well understood. We developed a method for the structural assessment of proteins in the foam state using hydrogen/deuterium exchange mass spectrometry. Adsorption sites and structural changes in human serum albumin (HSA) were identified in situ at the peptide-level resolution. The N-terminus and the loop (E492-T506), which contains hydrophobic amino acids, were identified as adsorption sites. Both the structural flexibility and hydrophobicity were considered to be critical factors for the adsorption of HSA at the AWI. Structural changes in HSA were observed after more than one minute of foaming and were spread widely throughout the structure. These structural changes at the foam AWI were reversible.

Human Serum Albumin in the Presence of Small Platinum Nanoparticles

Noble metal materials, especially platinum nanoparticles (Pt NPs), have immense potential in nanomedicine as therapeutic agents on account of their high electron density and their high surface area. Intravenous injection is proposed as the best mode to deliver the product to patients. However, our understanding of the reaction of nanoparticles with blood components, especially proteins, is far behind the explosive development of these agents. Using synchrotron radiation circular dichroism (SRCD), we investigated the structural and stability changes of human serum albumin (HSA) upon interaction with PEG-OH coated Pt NPs at nanomolar concentrations, conditions potentially encountered for intravenous injection. There is no strong complexation found between HSA and Pt NPs. However, for the highest molar ratio of NP:HSA of 1:1, an increase of 18 °C in the thermal unfolding of HSA was observed, which is attributed to increased thermal stability of HSA generated by preferential hydration. This work proposes a new and fast method to probe the potential toxicity of nanoparticles intended for clinical use with intravenous injection.

Research on the Interaction Mechanism and Structural Changes in Human Serum Albumin with Hispidin Using Spectroscopy and Molecular Docking.

The interaction between human serum albumin (HSA) and hispidin, a polyketide abundantly present in both edible and therapeutic mushrooms, was explored through multispectral methods, hydrophobic probe assays, location competition trials, and molecular docking simulations. The results of fluorescence quenching analysis showed that hispidin quenched the fluorescence of HSA by binding to it via a static mechanism. The binding of hispidin and HSA was validated further by synchronous fluorescence, three-dimensional fluorescence, and UV/vis spectroscopy analysis. The apparent binding constant (Ka) at different temperatures, the binding site number (n), the quenching constants (Ksv), the dimolecular quenching rate constants (Kq), and the thermodynamic parameters (∆G, ∆H, and ∆S) were calculated. Among these parameters, ∆H and ∆S were determined to be 98.75 kJ/mol and 426.29 J/(mol·K), respectively, both exhibiting positive values. This observation suggested a predominant contribution of hydrophobic forces in the interaction between hispidin and HSA. By employing detergents (SDS and urea) and hydrophobic probes (ANS), it became feasible to quantify alterations in Ka and surface hydrophobicity, respectively. These measurements confirmed the pivotal role of hydrophobic forces in steering the interaction between hispidin and HSA. Site competition experiments showed that there was an interaction between hispidin and HSA molecules at site I, which situates the IIA domains of HSA, which was further confirmed by the molecular docking simulation.

Deciphering conformational changes in human serum albumin induced by bile salts using spectroscopic and molecular modeling approaches

Bile salts are physiologically-important natural amphiphilic biosurfactants synthesized in the liver and play a vital role in the solubilization and digestion of dietary lipids, cholesterol, and other fat-soluble compounds in the body. Bile salts also exhibit pharmacological and biological uses as carriers for transporting scarce water-soluble drugs and other chemicals owing to their unique emulsifying, solubilizing capacities and micelle forming ability. In this context, we present an endeavor towards the possibilities underlying the interaction and binding between the most abundant plasma protein namely, human serum albumin (HSA) and bile salts to demonstrate an intriguing interplay of hydrophobic, electrostatic and hydrogen bonding effects using steady state absorption, fluorescence emission, anisotropy, time-resolved emission, and molecular modelling approaches. The outcome illuminates how amphiphilic interfaces of bile salts under various physico-chemical conditions trigger the conformational changes and binding affinities of native and molten-globule forms of HSA. To elucidate non-covalent interactions during HSA-bile salt supramolecular host–guest complex formation, changes in intrinsic (tryptophan) and extrinsic (ANS) fluorescence have been investigated. The results reveal upon binding of bile salts in subdomain IIA of HSA, the protein undergoes conformational changes mediated primarily by hydrophobic interactions. Furthermore, time-resolved fluorescence measurements provide important structural and dynamical insights into the protein-bile salt supramolecular complexes. Additionally, molecular docking studies on these complexes clearly reveal spontaneous binding of bile salts into subdomain IIA of HSA while suggesting that the binding affinity decreases with the decreasing order of hydrophobicity of the bile salts (NaDC > NaC > NaTC) (ΔGdock = −29.64 kJ mol−1, −26.15 kJ mol−1, −14.35 kJ mol−1), respectively. This study exclusively highlights the molecular mechanism of conformational perturbation in the native (pH 7) and molten-globule (pH 3) forms of HSA, induced by bile salts. We believe that the results reported herein will be helpful in the design and formulation of protein-bile salt-based pharmacological carriers suitable for drug delivery.

Simultaneous binding characterization of different chromium speciation to serum albumin.

The binding process between three species of chromium and serum albumin (SA) was investigated, as well as the interaction between K2Cr2O7 and bovine serum albumin (BSA) under coexistence of different chromium forms. CrCl3, K2Cr2O7 and Crpic bound to SA spontaneously through Van der Waals force, and their binding constants were 103-104M-1 at 298K, respectively. K2Cr2O7 and Crpic both had strong binding affinity for BSA, and significantly affected the secondary structure of BSA and the microenvironment surrounding amino acid residues. Chromium exhibited a greater fluorescence quenching constant towards HSA than toward BSA, and K2Cr2O7 induced greater conformational changes in human serum albumin (HSA) than in BSA. A weak binding of CrCl3 to BSA had no significant effect on the binding affinity of K2Cr2O7 to BSA. K2Cr2O7 and BSA have a greater binding affinity when coexisting with Crpic, and K2Cr2O7 induces a greater conformational change in BSA.

Understanding Conformational Changes in Human Serum Albumin and Its Interactions with Gold Nanorods: Do Flexible Regions Play a Role in Corona Formation?

The importance of protein-nanoparticle (NP) conjugates for biomedical applications has seen an exponential growth in the past few years. The protein corona formation on NPs with human serum albumin (HSA), being the most abundant protein in blood serum, has become one of the most studied protein analyses under NP-protein interactions as HSA is readily adsorbed on the surface of the NPs. Understanding the fate of the NPs in physiological media along with the change in biological responses due to the formation of the protein corona thus becomes important. We analyzed the HSA protein corona formation on gold nanorods (AuNRs) through different spectroscopic studies in addition to the effects of change in the protein concentration on the protein-NP interactions. Different imaging techniques such as high-resolution transmission electron microscopy, field emission scanning electron microscopy, and atomic force microscopy were used to determine the morphology and the dimensions of the nanorods and the protein-nanorod conjugates. Fourier-transform infrared data showed a reduction in the α-helix content and an increase in β-sheet content for the HSA-AuNR conjugate compared to the native protein. A decrease in steady-state fluorescence intensity occurred with instant addition of AuNR to HSA showing better and efficient quenching of Trp fluorescence for the lower concentration of protein. Time-correlated single photon counting results showed greater energy transfer efficiency and faster decay rate for higher concentrations of proteins. The circular dichroism study gives insight into the secondary structural changes due to unfolding, and a greater change was observed for lower concentrations of protein due to a thermodynamically stable protein corona formation. Surface-enhanced Raman spectroscopy (SERS) indicated the presence of aromatic residues such as Phe, Tyr, and Cys that appear to be close to the surface of the AuNRs in addition to hydrophobic interactions between AuNR and the protein. The disordered and flexible regions mapped onto HSA (PDB: 1AO6), predicted by the intrinsically disordered region predictors, point toward the interactions of similar residues with the nanorods observed from SERS and fluorescence studies. These studies could provide a clearer understanding of the interactions between HSA and AuNRs for possible biological applications.

Investigation of structural changes in human serum albumin after binding with elaidic acid

Elaidic acid (EA) is an unsaturated trans-fatty acid and one of the most important fatty acids obtained from solidifying vegetable oils. The increase in the consumption of EA is associated with rise and decline of LDL-cholesterol and HDL-cholesterol levels, respectively. It activates pro-inflammatory markers in the body and increases heart stroke and cardiovascular diseases. In this study, to understand the impact of EA in humans, we investigated the molecular interaction between EA and human serum albumin (HSA) and its binding affinity using spectroscopic, molecular dynamics (MD), and molecular docking techniques. According to fluorescence quenching experiments, EA could quench the inherent fluorescence of HSA via a static quenching mechanism. The thermodynamic characteristics of EA binding on HSA demonstrated hydrogen bonds and Van der Waals forces within the interaction, which matched molecular docking results. Further, the negative amount of ΔG° indicated the spontaneous binding of HSA to EA. Based on Förster's resonance energy transfer, the spatial distance between EA and HSA was 3.493 nm. The far-UV circular dichroism spectrum showed the decrease in the contents of the β-sheet and α-helix in the HSA structure. According to the RMSF data, the HSA structure's flexibility was diminished. The molecular dynamic simulation and spectroscopic studies also indicated that the binding interactions altered the milieu of the Trp chromophore and HSA structures. All experimental data could be matched with molecular dynamics simulation.

Methylglyoxal-induces multiple stable changes in human serum albumin before forming nephrotoxic advanced glycation end-products: Injury demonstration in human embryonic kidney cells

The minor fraction of methylglyoxal that is not metabolized in healthy humans reacts with macromolecules to form AGEs. In diabetics, the formation of MG is accelerated; its level may be enhanced multifold. The glyoxalase enzymes responsible for the regular and effective clearance of excess methylglyoxal may become defective in diabetes mellitus leading to its retention in cells and plasma. The methylglyoxal-modified-HSA was prepared, characterised by multiple biophysical techniques and biochemical (s) and its damaging effect was examined on embryonic kidney cell line HEK 293. The UV results showed hyperchromicity in MG-modified-HSA while nitroblue tetrazolium and fluorescence data suggested AGEs formation in comparison to control HSA. Upward shift of negative peaks in CD suggested reduction in α-helicity. Accelerated mobility and diffused broad bands observed in native and SDS polyacrylamide gel, respectively suggest neutralization of some of the positive charges on MG-modified-HSA as well as generation of cross-links. As observed by trypan blue assay, MTT, LDH activity assay, acridine orange, propidium iodide, ethidium bromide, 4′,6-diamidino-2-phenylindole (DAPI) staining and ROS measurements, the MG-HSA AGEs caused damage to human embryonic kidney cells. The data suggest that MG-HSA AGEs may trigger powerful inflammatory responses at cellular level which might set the stage for nephrotoxicity in diabetics.

Investigation of the binding properties of 3,4-dihydroxybenzaldehyde from Salvia miltiorrhiza (Bunge) with human serum albumin via multi-spectroscopic and molecular docking techniques

To investigate the binding properties of 3,4-dihydroxybenzaldehyde with human serum albumin, as well as the structural changes of human serum albumin under a simulated physiological pH value (a pH of 7.4) and a high 3,4-dihydroxybenzaldehyde concentration, a series of techniques, i.e., fluorescence, synchronous fluorescence, ultraviolet-visible absorption, Fourier-transform infrared spectroscopy, and molecular docking simulation, were employed. Steady state fluorescence showed that 3,4-dihydroxybenzaldehyde quenched the intrinsic fluorescence of human serum albumin via a static mechanism. The 3,4-dihydroxybenzaldehyde-human serum albumin complex had a strong affinity (Kb = 105 M-1) at various temperatures. It was shown that 3,4-dihydroxybenzaldehyde was bound to the IB subdomain of human serum albumin primarily via hydrogen bonding and van der Waals forces at high 3,4-dihydroxybenzaldehyde concentrations, based on the results of the thermodynamic and molecular docking. Furthermore, the fluorescence emission spectrum and Fourier-transform infrared spectroscopy results indicated that the binding distance between 3,4-dihydroxybenzaldehyde and human serum albumin was 4.42 nm. In addition, 3,4-dihydroxybenzaldehyde induced conformational changes of human serum albumin. These findings provide reasonable evidence for further understanding the distribution of 3,4-dihydroxybenzaldehyde when it spreads into human blood serum, which may be helpful in food and medicine research.

Changes in Glycated Human Serum Albumin Binding Affinity for Losartan in the Presence of Fatty Acids In Vitro Spectroscopic Analysis.

Conformational changes in human serum albumin due to numerous modifications that affect its stability and biological activity should be constantly monitored, especially in elderly patients and those suffering from chronic diseases (which include diabetes, obesity, and hypertension). The main goal of this study was to evaluate the effect of a mixture of fatty acids (FA) on the affinity of losartan (LOS, an angiotensin II receptor (AT1) blocker used in hypertension, a first-line treatment with coexisting diabetes) for glycated albumin—simulating the state of diabetes in the body. Individual fatty acid mixtures corresponded to the FA content in the physiological state and in various clinical states proceeding with increased concentrations of saturated (FAS) and unsaturated (FAUS) acids. Based on fluorescence studies, we conclude that LOS interacts with glycated human serum albumin (af)gHSA in the absence and in the presence of fatty acids ((af)gHSAphys, (af)gHSA4S, (af)gHSA8S, (af)gHSA4US, and (af)gHSA8US) and quenches the albumin fluorescence intensity via a static quenching mechanism. LOS not only binds to its specific binding sites in albumins but also non-specifically interacts with the hydrophobic fragments of its surface. Incorrect contents of fatty acids in the body affect the drug pharmacokinetics. A higher concentration of both FAS and FAUS acids in glycated albumin reduces the stability of the complex formed with losartan. The systematic study of FA and albumin interactions using an experimental model mimicking pathological conditions in the body may result in new tools for personalized pharmacotherapy.

Spectroscopic and molecular docking studies for characterizing binding mechanism and conformational changes of human serum albumin upon interaction with Telmisartan

Human serum albumin (HSA), one of the most copious plasma proteins is responsible for binding and transportation of many exogenous and endogenous ligands including drugs. In this study, we intended to explore the extent and types of binding interaction present between HSA and the antihypertensive drug, telmisartan (TLM). The conformational changes in HSA due to this binding were also studied using different spectroscopic and molecular docking techniques. The spectral shifting and intensity variations upon interaction with TLM were studied using FT-IR spectroscopy. Binding constant and the change in absorption of HSA at its λmax was analyzed using absorption spectroscopy. Eventually, the types and extent of binding interactions were confirmed using molecular docking technique. Results have shown that TLM significantly interacts with the binding site-1 of HSA utilizing strong hydrogen bonding with Glu292, and Lys195 residues. The UV-absorption intensities were found to be decreased serially as the drug concentration increased with a binding constant of 1.01 × 103 M−1. The secondary structure analysis using FT-IR spectroscopy also revealed a marked reduction in the α-helix (56%) component of HSA on interaction. This study gives critical insights into the interaction of TLM with HSA protein which eventually affects the concentration of TLM reaching the site of action and ultimately its therapeutic profile.

FTIR Spectroscopy Study of the Secondary Structure Changes in Human Serum Albumin and Trypsin under Neutral Salts.

The effect of neutral salts on protein conformation was first analyzed by Hofmeister in 1888, however, even today this phenomenon is not completely understood. To clarify this effect, we studied changes in the secondary structure of two proteins: human serum albumin with predominantly α-helical structure and porcine pancreas β-trypsin with the typical β-structural arrangement in aqueous solutions of neutral salts (KSCN, KCl, (NH4)2SO4). The changes in the secondary structure were studied at 23 °C and 80 °C by using the second derivative deconvolution method of the IR spectra. Our results demonstrated that the ability of the salts to stabilize/destabilize these two proteins correlates with the Hofmeister series of ions. At the same time, some exceptions were also observed. The destabilization of the native structures of both α-helical albumin and β-structural trypsin upon interaction with neutral salts leads to the formation of intermolecular β-sheets typical for amyloid fibrils or amorphous aggregates. Thus, our quantitative FTIR-spectroscopy analysis allowed us to further clarify the mechanisms and complexity of the neutral salt actions on protein structures which may lead to strategies preventing unwelcome misfolding of proteins.

Preliminary study of human serum albumin level in early warning onset of preeclampsia

Objective: To observe the dynamic changes of human serum albumin (HSA) level during pregnancy and study the early warning significance of HSA level on the onset of preeclampsia (PE) . Methods: Totally 369 PE pregnant women (PE group) and 309 normal pregnant women (control group) without PE who admitted in Haidian Maternal and Child Health Hospital from January 2013 to December 2017 were selected. HSA levels were tested before meeting the criterion of PE in the first trimester, the early-third trimester and the late-third trimester, the difference between the two groups were compared. The relationship between the HSA level and the incidence of complications in PE patients was analyzed. Results: (1)The mean values of HSA level in PE group and control group were (41.9±3.1) versus (40.0±2.2) g/L, (34.2±2.7) versus (35.4±2.7) g/L and (33.7±2.9) versus (36.7±3.3) g/L in the first trimester,the early-third trimester and the late-third trimester respectively,the difference in the first trimester was no significance (P>0.05), while the differences in the early-third trimester and the late-third trimester were both significant (all P<0.05). (2) The HSA level during pregnancy of PE group showed a continuous downward trend, while the control group was V-shaped trend. The receiver operating characteristic (ROC) curve analysis showed that PE could be early warned by the decrease of HSA level in PE group [area under curve (AUC)=0.742, cut-off value=5.97 g/L, sensitivity 70.8%, specificity 62.8%], the same result was in severe PE (AUC=0.756, cut-off value=6.85 g/L, sensitivity 70.8%, specificity 72.0%). The level of HSA was negatively correlated with the incidence of complications (r=-0.19, P<0.01). Conclusions: Excessive decrease of HSA level is an early warning factor for PE onset. The higher the baseline of HSA level and the greater the extent of pregnancy decline, the risk of PE in pregnant women is higher. The lower of HSA level in PE, the incidence of complications is higher. The excessive decrease of HSA level may be the first clinical manifestation before the onset of clinical symptoms of PE, so it may be the warning factor and one of the laboratory indicators in the PE sub-clinical stage.

Quantitative Photo-crosslinking Mass Spectrometry Revealing Protein Structure Response to Environmental Changes.

Protein structures respond to changes in their chemical and physical environment. However, studying such conformational changes is notoriously difficult, as many structural biology techniques are also affected by these parameters. Here, the use of photo-crosslinking, coupled with quantitative crosslinking mass spectrometry (QCLMS), offers an opportunity, since the reactivity of photo-crosslinkers is unaffected by changes in environmental parameters. In this study, we introduce a workflow combining photo-crosslinking using sulfosuccinimidyl 4,4′-azipentanoate (sulfo-SDA) with our recently developed data-independent acquisition (DIA)-QCLMS. This novel photo-DIA-QCLMS approach is then used to quantify pH-dependent conformational changes in human serum albumin (HSA) and cytochrome C by monitoring crosslink abundances as a function of pH. Both proteins show pH-dependent conformational changes resulting in acidic and alkaline transitions. 93% and 95% of unique residue pairs (URP) were quantifiable across triplicates for HSA and cytochrome C, respectively. Abundance changes of URPs and hence conformational changes of both proteins were visualized using hierarchical clustering. For HSA we distinguished the N–F and the N–B form from the native conformation. In addition, we observed for cytochrome C acidic and basic conformations. In conclusion, our photo-DIA-QCLMS approach distinguished pH-dependent conformers of both proteins.

Immunoreactivity changes of human serum albumin and alpha-1-microglobulin induced by their interaction with dendrimers.

Dendrimers are hyperbranched polymers for delivery of therapeutic genetic material to cancer cells. The fine tuning chemical modifications of dendrimers allow for the modification of the composition. The architecture and the properties of dendrimers are key factors to improve their in vitro and in vivo properties such as biocompatibility with cells and tissues and their pharmacokinetic/pharmacodynamic behavior. The side effects of dendrimers on structure and function of proteins is an important question that must be addressed. We herein describe the effect of newly synthesized piperidine-based cationic phosphorous dendrimers of 2 generations and commercial cationic, neutral and anionic poly(amidoamine) (PAMAM) dendrimers of 4th generation on immunochemical properties of 2 serum proteins: human serum albumin (HSA) and alpha-1-microglobulin (A1M). Both can bind and transfer ligands in blood, including hormones, fatty acids, toxins and drugs, and have immunoreactivity properties. Comparing the effects of piperidinium-terminated phosphorus and cationic, neutral and anionic PAMAM dendrimers on HSA and A1M, we conclude that, in the case of equimolar complexes, these dendrimers had no significant effect on immunoreactivity of proteins. In contrast, the formation of complexes in which a protein is fully bound to dendrimers leads to partial (1.2-2.3 times) reduction in protein immunoreactivity. The most important fact is that dendrimer-induced change in immunoreactivity of proteins is not complete, even if the protein is entirely bound by dendrimers. This means that the application of dendrimers in vivo will not totally hamper the immunoreactivity of these proteins and antibodies.

Interaction between human serum albumin and toxic free InP/ZnS QDs using multi-spectroscopic study: An excellent alternate to heavy metal based QDs

Heavy metal free and non-toxic semiconductor nanocrystals are the most interested segment of the current bio-nanotechnology. In the present paper, structural changes of human serum albumin (HSA) upon its interaction with InP/ZnS QDs coated with amphilic poly ethylene glycol was probed by employing multi-spectroscopic tools. Steady state and time resolved fluorescence studies implied formation of non-fluorescent complex between HSA and QDs resulting out of static quenching. Interestingly, temperature dependent fluorescence quantitative studies show that at higher temperatures these bioconjugates are unstable and static quenching mechanism turns towards dynamic quenching as QDs diffuses through excited state of HSA. Binding and synchronous fluorescence spectroscopic analysis show that QDs binds predominantly to Trp-214 residue in the HSA subdomain IIA with hydrophobic forces. Hence, observed blue shift of about 5 nm in PL spectra of HSA is the clear sign of conformational deformation as the Trp-214 residue gradually enters into hydrophobic microenvironment during its conjugation with QDs. Energy transfer efficiency (ET), transfer rate (KT) and critical transfer distance (R0) were all calculated for HSA and PEG-InP/ZnS QDs system from Förster resonance energy transfer (FRET) studies. Gibb's free energy, enthalpy and entropy obtained from temperature dependent fluorescence studies implied, HSA-InP/ZnS QDs bioconjugation is spontaneous, endothermic, entropy driven and is in accordance with the second law of thermodynamics. Hill coefficient (n ≅ 1) and binding affinities (kb) confirmed strong binding of HSA to QDs surface through ‘non-cooperative interactions’. Also, experimental result of FTIR spectra confirmed the adsorption of HSA onto the surfaces of PEG-InP/ZnS QDs. In addition these results proves that greener, heavy metal free and non-toxic semiconductor nanoparticle (PEG-InP/ZnS QDs) can potentially replace heavy metal and toxic QDs in biophysical fields like conformational studies of proteins.

Maslinic acid conjugate with 7-amino-4-methylcoumarin as probe to monitor the temperature dependent conformational changes of human serum albumin by FRET.

Synthesis, characterization and spectroscopic investigation of maslinic acid labeled with fluorescent 7-amino-4-methylcoumarin is reported. It was found that the coumarin-maslinic derivative (MaCo) forms an excellent fluorescence resonance energy transfer (FRET) pair with the tryptophan (Trp) residue of human serum albumin (HSA). This feature allowed for monitoring HSA conformational alterations by measuring the distance between donor (Trp) and acceptor (MaCo) through Förster energy transfer mechanism. Displacement experiments confirmed that MaCo binds to subdomain IIA of HSA with independence of temperature. It was observed that, in the temperature range 35-45 °C, the fluorescence emission maximum of HSA-MaCo complex decreased, whereas in the range 45 °C-65 °C, an increment was detected. The concomitant change in the polarity of environment surrounding Trp was confirmed by red edge excitation shift experiments. Thermal denaturation of HSA was followed by time-resolved fluorescence spectroscopy. Average lifetime of Trp residue decreased with temperature due to the increment of solvent collisions and changes in the solvent exposure of Trp. To discriminate the importance of each effect, lifetime of N-Acetyl-L-tryptophanamide (NATA) at different temperatures was measured. Circular dichroism (CD) studies confirmed the loss of secondary structure of HSA with increasing temperature and showed a different trend in the conformational transformation below and above 45 °C, in agreement with steady-state and time-resolved fluorescence experiments.

Characterization of the structural changes of human serum albumin upon interaction with single-walled and multi-walled carbon nanotubes: spectroscopic and molecular modeling approaches

Through the incorporation of spectorescopic and molecular methods of modeling, the researchers investigated the interaction between Carbon Nanotubes (CNTs) and Human Serum Albumin (HSA). Fluorescence spectroscopy revealed the ability in both single-wall and multi-wall CNTs to quench the spectrum through a static quenching procedure obtained from the Stern–Volmer quenching constant (Ksv) at three different temperatures. The Ksv values of HSA–CNTs complexes were 1.96 × 105 M−1 and 2.44 × 105 M−1 that showed two different behaviors of interaction between HSA and CNTs. The Van’t Hoff equation was used to calculate thermodynamic parameters of Gibbs free energy (ΔG°), entropy (ΔS°) and enthalpy changes (ΔH°). The binding distances (r) between the donor (Trp residue of HSA) and acceptor (CNTs) was measured through the Forster theory of non-radiative energy transfer, and it was found to be less than 7 nm. The conformational changes of protein in the presence of CNTs were revealed through synchronous fluorescence spectra and three-dimensional fluorescence spectra analysis. The experimental results were confirmed through molecular modeling technique. Circular dichroism technique showed the secondary structure changes of HSA upon interaction with CNTs. The complex formation of HSA and CNTs was determined by the measurement of the electric conductivity. The molecular modeling technique determined the binding site of CNTs that were embedded in the subdomain IIIB of HSA. These analyses play a major role in drug delivery and pharmacodynamics studies for better understanding of nanotechnology levels.

Biomolecular interactions of amphotericin B nanomicelles with serum albumins: A combined biophysical and molecular docking approach

In this work, we investigated the interaction of amphotericin B (AmB) nanomicelles on the binding affinity and conformational change of human serum albumin (HSA) in comparison with bovine serum albumin (BSA) under physiological conditions by conducting several spectroscopic techniques further confirmed through molecular docking approaches. The experimental results showed that AmB nanomicelles could bind to both HSA and BSA to form protein/drug complexes with one binding site, and the binding process was spontaneous under physiological conditions. Fluorescence studies revealed that the quenching mechanism of these complexes was static quenching rather than dynamic quenching and exhibited strong binding between serum albumin and AmB nanomicelles. The results from UV–Visible spectra, FT-IR spectra, and CD spectra revealed that the AmB formulations affected the structure of both HSA and BSA proteins by changing the microenvironment around the tryptophan residues of protein and caused a secondary structure change of the protein with the loss of helical stability. The molecular docking experiments also supported the above results and effectively proved the binding and changes in the conformation of serum albumins by AmB micelles. This finding provides information of in vitro drug-plasma protein interactions for further study on the AmB binding mechanism and the pharmacodynamics and pharmacokinetics.