Secondary Structure Of Bovine Serum Albumin Research Articles

In vitro DNA and BSA-binding, cell imaging and anticancer activity against human carcinoma cell lines of mixed ligand copper(II) complexes

Binding studies of two water soluble copper(II) complexes of the type [Cu(phen-dion)(diimine)Cl]Cl, where phen-dione is 1,10-phenanthroline-5,6-dione and diimine is 1,10-phenanthroline (1) and 2,2′-bipyridine (2), with fish sperm DNA (FS-DNA) and bovine serum albumin (BSA) have been examined under physiological conditions by a series of experimental methods (UV–Vis absorption, fluorescence, viscosity, cyclic voltammetry (CV) and circular dichroism (CD) spectroscopic techniques). The experimental results indicate that the complexes interact with FS-DNA by electrostatic and partial insertion of pyridyl rings between the base stacks of double-stranded DNA. The complexes could quench the intrinsic fluorescence of BSA with the binding constants (Kbin) of 32×105M−1 (1) and 1.7×105M−1 (2) at 290K. The quenching mechanism, thermodynamic parameters, the number of binding sites and the effect of the Cu(II) complexes on the secondary structure of BSA have been explored. The in vitro anticancer chemotherapeutic potential of two copper(II) complexes against the three human carcinoma cell lines (MCF-7, A-549, and HT-29) and one normal cell line (DPSC) were evaluated by MTT assay. The results of in vitro cytotoxicity indicate that the complex (1) has greater cytotoxicity activity against all of the cell lines, especially HT-29 with IC50 values of 1.8μM. Based on the IC50 values, these complexes did not display an apparent cyto-selective profile, because it would appear that two complexes are toxic to all four model cell lines. The microscopic analyses of the cancer cells confirm results of cytotoxicity.

Study on the interaction between bovine serum albumin and starch nanoparticles prepared by isoamylolysis and recrystallization

The current study primarily investigated the interaction of bovine serum albumin (BSA) with starch nanoparticles (SNPs) prepared by isoamylolysis and recrystallization using UV–vis, fluorescence, transmission electron microscopy (TEM), Fourier transform infrared (FTIR) and circular dichroism (CD). The enhanced absorbance observed by UV–vis spectroscopy and decreased intensity of fluorescence spectroscopy suggested that BSA could bind to SNPs and form a BSA–SNP complex. The synchronous fluorescence spectra revealed that the emission maximum of Tyr residue (at Δλ=15nm) was red-shifted at the investigated concentrations range, indicating that the conformation of BSA was changed. Quenching parameters showed that the quenching effect of SNPs was static quenching. TEM images showed that the SNPs were surrounded by protein coronae, indicating that nanoparticle–protein complexes had formed. The FTIR and CD characterization indicated that the SNPs induced structural changes in the secondary structure of BSA.

Systematic investigation of the toxic mechanism of PFOA and PFOS on bovine serum albumin by spectroscopic and molecular modeling

Perfluorinated compounds (PFCs), an emerging class of globally environmental contaminations, pose a great threat to humans with wide exposure from food and other potential sources. The effects of perfluorooctane sulfonate (PFOS) and perfluorooctanoic acid (PFOA) on bovine serum albumin (BSA) under normal physiological conditions were characterized by fluorescence, UV–Vis absorption, Fourier transform infrared (FT-IR) spectroscopy and molecular docking methods. The fluorescence study suggested that the fluorescence quenching of BSA by PFCs was a static procedure forming a PFCs–BSA complex. The negative values of enthalpy change (ΔH) and entropy change (ΔS) indicated that van der Waals forces and hydrogen bonds were the dominant intermolecular forces in the binding of PFCs to BSA. The displacement experiments of site markers and molecular docking revealed that the binding of PFOA to BSA took place in sub-domain IIA (Sudlow site I) whereas PFOS was mainly located in the sub-domain IIIA (Sudlow site II) and partially bound into site I. Furthermore, the results of UV–Vis and FT-IR spectra demonstrated that the microenvironment and the secondary structure of BSA were changed in the presence of PFCs. These results indicated that PFCs indeed impact the conformation of BSA and PFOS was more toxic than PFOA, which were supported by theoretical molecular modeling methods.

Mechanistic and conformational studies on the interaction of a platinum(II) complex containing an antiepileptic drug, levetiracetam, with bovine serum albumin by optical spectroscopic techniques in aqueous solution.

Fluorescence spectroscopy in combination with circular dichroism (CD) and ultraviolet-visible (UV-vis) absorption spectroscopy were employed to investigate the binding of a new platinum(II) complex containing an antiepileptic drug "Levetiracetam" to bovine serum albumin (BSA) under the physiological conditions. In the mechanism discussion, it was proved that the fluorescence quenching of BSA by Pt(II) complex is a result of the formation of Pt(II) complex-BSA complex. The thermodynamic parameters ΔG, ΔH, and ΔS at different temperatures (283, 298, and 310 K) were calculated, and the negative value for ΔH and ΔS indicate that the hydrogen bonds and van der Waals interactions play major roles in Pt(II) complex-BSA association. Binding studies concerning the number of binding sites (n~1) and apparent binding constant K b were performed by fluorescence quenching method. The site marker competitive experiments indicated that the binding of Pt(II) complex to BSA primarily took place in site II. Based on the Förster's theory, the average binding distance between Pt(II) complex and BSA was obtained (r = 5.29 nm). Furthermore, UV-vis, CD, and synchronous fluorescence spectrum were used to investigate the structural change of BSA molecules with addition of Pt(II) complex. These results indicate that the binding of Pt(II) complex to BSA causes apparent change in the secondary structure of BSA and do affect the microenvironment around the tryptophan residue.

BSA/polyelectrolyte core–shell nanoparticles for controlled release of encapsulated ibuprofen

Bovine serum albumin (BSA) based core–shell nanoparticles were developed as carrier systems for drug transportation. At pH=3, the oppositely charged polyelectrolytes: poly(sodium-4-styrene)sulphonate (PSS) and the chitosan (Chit) bind to the positively charged protein via electrostatic interactions. We applied ibuprofen (IBU) as model molecule which has low solubility. The changes in the BSA's secondary structure during the steps of the synthesis were inspected by FT-IR measurements. The size and the zeta potential were determined by dynamic light scattering (DLS). The changes in the structure and in the size were investigated by small angle X-ray scattering (SAXS) too, for each composite. The release of the ibuprofen was studied by vertical diffusion cell (Franz cell) at pH 7.4 at 25 and 37.5°C. The structure of the core–shell nanoparticles have significantly changed as the pH has risen from 3.0 to 7.4. Kinetic models were used to describe the release mechanism. The experimental results demonstrated that the BSA has an ordered structure at pH=3 which will become random coil by adding ibuprofen. The first shell restores the ordered structure of the protein. The controlled release was carried out; the IBU release decreased by 40% in the case of two-layered composites compared with the “naked” BSA.

Design and fabrication of a food-grade albumin-stabilized nanoemulsion

A food-grade biopolymer-emulsion delivery system was developed for improving or extending the functional performance of the bioactive components of food. Droplet size, rheological and interfacial measurements, scanning electron microscopy (SEM), thiobarbituric acid reactive substance (TBARS) assay and Fourier transform-infrared (FTIR) analysis were used to study the effect of sonication time and power, poly ethylene glycol (PEG) addition, heat treatment and oil fraction on physical and chemical stabilities of bovine serum albumin (BSA)-stabilized corn oil-in-water (O/W) emulsions prepared by ultrasonic emulsification. The findings showed that sonication power and time had significant effects on the reduction of mean droplet diameter to 75 nm in the presence of PEG. PEG acted as an enhancer of viscosity of the aqueous phase. PEG had negligible influence on the interfacial tension of the emulsion. An increase in the oil fraction of up to 20 wt% had no effect on the mean droplet diameter of BSA emulsion containing PEG. Mean droplet diameter of BSA emulsions that contained PEG 10000 and 300, increased to 170 and 250 nm, respectively, after 60 days of storage at 6 °C. FTIR analysis showed that the addition of PEG had no major influence on the secondary structure of BSA. The proportion of unadsorbed protein in the PEG-contained nanoemulsions was lower than that in emulsions without PEG. Based on TBARS content the oxidative stability of sonicated emulsions in the presence of PEG were less than other formulations.

Binding of a novel bacteriostatic agent—chitosan oligosaccharides–kojic acid graft copolymer to bovine serum albumin: spectroscopic and conformation investigations

In this paper, a novel chitosan oligosaccharides derivative—chitosan oligosaccharides–kojic acid graft copolymer (COS/KA), COS, and KA were selected to investigate the binding to bovine serum albumin (BSA) using UV–Vis spectroscopy, fluorescence, synchronous fluorescence, and circular dichroism (CD) at pH 7.4. The results indicated that the tryptophan residues of BSA to the distance of COS/KA, COS, and KA were 2.0 × 1010 L mol−1 s−1 for COS/KA–BSA, COS–BSA, and KA–BSA at 298, 302, 306, 310 K, respectively, those confirmed that the quenching process belongs to static quenching. The thermodynamic parameters ∆H°, ΔG°, ΔS° at different temperatures were calculated and indicated that hydrophobic and electrostatic forces played important roles in the binding process, and the enhanced binding affinity mainly associated with the increase of the hydrophobicity. Moreover, the CD data demonstrated that the secondary structure of BSA was slightly altered in the presence of COS/KA, COS, and KA, with different reduced α-helix contents (46.30 ± 0.20, 48.60 ± 0.40, 50.70 ± 0.20), respectively. The work provides basic data for the binding mechanism of COS/KA with BSA in vitro.

Interaction of norfloxacin with bovine serum albumin studied by different spectrometric methods; displacement studies, molecular modeling and chemometrics approaches

Serum albumins as major target proteins can bind to other ligands leading to alteration of their pharmacological properties. The mechanism of interaction between norfloxacin (NFLX) with bovine serum albumin (BSA) was investigated. Fuorescence quenching of serum albumin by this drug was found to be a static quenching process. The binding sites number, n, apparent binding constant, K, and thermodynamic parameters were calculated at different temperatures. The distance, r, between donor, BSA, and acceptor, NFLX, was calculated according to the Forster theory of non-radiation energy transfer. Also binding characteristics of NFLX with BSA together with its displacement from its binding site by kanamycin and effect of common metal ions on binding constant were investigated by the spectroscopic methods. The conformational change in the secondary structure of BSA upon interaction with NFLX was investigated qualitatively from synchronous fluorescence spectra, Fourier Transform Infrared (FTIR) and circular dichroism (CD) spectrometric methods. 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. The results showed that the conformation of BSA changed in the presence of NFLX. For the first time, displacement studies were used for this interaction; displacement studies showed that NFLX was displaced by phenylbutazon and ketoprofen but was not displaced by ibuprofen indicating that the binding site of NFLX on albumin was site I. In addition a powerful chemometrics method, multivariate curve resolution-alternating least square, was used for resolution of spectroscopic augmented data obtained in two different titration modes in order to extract spectral information regardless of spectral overlapping of components.

Exploring the interaction of a micelle entrapped biologically important proton transfer probe with the model transport protein bovine serum albumin.

This article describes the interaction of a micelle entrapped pharmaceutically important isoindole fused imidazole derivative, namely, 1-(2-hydroxy-5-methyl-phenyl)-3,5-dioxo-1H-imidazo-[3,4-b] isoindole (ADII), with the model transport protein bovine serum albumin (BSA). Different spectroscopic techniques such as steady state absorption, emission, circular dichroism, dynamic light scattering, etc., have been employed to explore preferential interaction of this drug template with micelles and protein BSA. Binding of ADII with BSA is found to be enormously modified when it is released from the micellar environment. The binding constant of the ADII-BSA complex is reduced when the probe is released from anionic SDS micelle, whereas the binding is observed to be strengthened in cationic CTAB micellar medium due to the formation of a 1:2 complex (ADII-BSA). Time-resolved studies also support our steady state findings that the released drug from the micellar environment is found to be strongly bound with the protein BSA. Circular dichroism (CD) and dynamic light scattering (DLS) study reveals that the secondary structure of BSA gets some stabilization in SDS medium after binding of drug template to protein. The probable binding location of the probe within the protein cavity (hydrophilic subdomain IA) has been explored from an AutoDock-based blind docking simulation study.

Light-Responsive Polyion Complex Micelles with Switchable Surface Charge for Efficient Protein Delivery.

In this letter, light-responsive protein-encapsulated polyion complex (PIC) micelles were prepared by self-assembly of cationic block copolymer poly(N,N-dimethyl-N-(2-(methacryloyloxy)ethyl)-N-((2-nitrobenzyl)oxy)-2-oxoethanaminium bromide)-block-poly(carboxybetaine methacrylate) (PDMNBMA-b-PCBMA) and negatively charged bovine serum albumin (BSA). The PIC micelles were well characterized by dynamic light scattering (DLS) and transmission electron microscopy (TEM). From the zeta potential measurement, the increase of the zeta potential of PIC micelles from ∼10 to ∼20 mV was observed when the solution pH decreased from 7.4 to 6.5, which could enhance the intracellular protein delivery efficiency. Moreover, the positively charged PDMNBMA blocks can be transformed to zwitterionic carboxybetaine units under UV irradiation, which could result in the disassembly of the PIC micelles. The release of BSA can therefore be drastically accelerated in the presence of UV irradiation. Meanwhile, the circular dichroism (CD) spectroscopy confirmed that the secondary structure of BSA was unaffected during the UV irradiation process.

Secondary structure investigation of bovine serum albumin (BSA) by Fourier transform infrared (FTIR) spectroscopy in the amide III region

Fourier transform infrared spectroscopy is widely used to analyze protein secondary structures. The common strategy in this field is to analyze the conformation sensitive 1700-1600 cm -1 amide I region of protein FTIR spectrum. Though the amide III region of protein is also sensitive to secondary structural changes, its potential for protein secondary structural analysis is largely unexplored. In this paper, we performed a detailed investigation on the second structural analysis of bovine serum albumin by monitoring the spectral variation of the amide III band under a variety of pH conditions by FTIR spectroscopy and FTIR second derivative spectroscopy. Our results show that both acidic and basic conditions have pronounced effects on the overall secondary structures of BSA, suggesting denaturation effects. Furthermore, we observe that the amide III band profiles under acidic and basic conditions appear to be quite different. Our results clearly demonstrate that the amide III region is a promising probing region for protein secondary structural analysis.

Spectroscopy and docking simulations of the interaction between lochnericine and bovine serum albumin.

Lochnericine (LOC) is a component of Voacanga africana, which is a type of traditional medical food in Africa widely used for treating diseases. In this article, the interaction between LOC and bovine serum albumin (BSA) was studied by fluorescence spectroscopy. Furthermore, Fourier transform infrared (FTIR), Raman and circular dichroism (CD) were used to investigate the structural changes of BSA. The experimental results consistently indicated that LOC changed the secondary structure of BSA. Three structure-similar components were used to study the interference experiments. The molecular modeling results showed that LOC could bind within not only sites I and II, but also bind the cavity of subdomain IB.

MicroglassificationTM: A Novel Technique for Protein Dehydration

The dehydration of biologics is commonly employed to achieve solid-dose formulation and enhanced stability during long-term preservation. We have developed a novel process, Microglassification™, which can rapidly and controllably dehydrate protein solutions into solid amorphous microspheres at room temperature. Single bovine serum albumin (BSA) microdroplets were suspended in pentanol or decanol using a micropipette, and the dynamic changes in droplet dissolution were observed in real-time and correlated to protein's water of hydration, medium's water activity, and microsphere protein concentration. Microglassification™ was also carried out at bulk scale, and changes in BSA secondary structure were analyzed by Fourier transform infrared spectroscopy and fluorescence spectroscopy; multimer formation was detected by native gel electrophoresis. BSA concentration in the microsphere increased with solvent exposure time and decreasing water activity. Image analysis at single particle and bulk scale showed the formation of solid BSA microspheres with a maximum protein concentration of 1147 ± 32 mg/mL. The native BSA samples were dehydrated to approximately 450 waters per BSA, which is well below monolayer coverage of 1282 waters per BSA. The secondary structure of Microglassified™ BSA reverted to native-like conformation upon rehydration with only minor irreversible aggregation (2.7%). Results of the study establish the efficacy of the Microglassification™ for the successful dehydration of biologics.

Detection of interaction between lysionotin and bovine serum albumin using spectroscopic techniques combined with molecular modeling

A combination of fluorescence, UV-Vis absorption, circular dichroism (CD), Fourier transform infrared (FT-IR) and molecular modeling approaches were employed to determine the interaction between lysionotin and bovine serum albumin (BSA) at physiological pH. The fluorescence titration suggested that the fluorescence quenching of BSA by lysionotin was a static procedure. The binding constant at 298K was in the order of 10(5)Lmol(-1), indicating that a high affinity existed between lysionotin and BSA. The thermodynamic parameters obtained at different temperatures (292, 298, 304 and 310K) showed that the binding process was primarily driven by hydrogen bond and van der Waals forces, as the values of the enthalpy change (ΔH°) and entropy change (ΔS°) were found to be -40.81±0.08kJmol(-1) and -35.93±0.27Jmol(-1)K(-1), respectively. The surface hydrophobicity of BSA increased upon interaction with lysionotin. The site markers competitive experiments revealed that the binding site of lysionotin was in the sub-domain IIA (site I) of BSA. Furthermore, the molecular docking results corroborated the binding site and clarified the specific binding mode. The results of UV-Vis absorption, CD and FT-IR spectra demonstrated that the secondary structure of BSA was altered in the presence of lysionotin.

The Spectroscopy Study of the Binding of an Active Ingredient of Dioscorea Species with Bovine Serum Albumin with or without Co(2+) or Zn(2+).

Diosgenin (DIO) is the active ingredient of Dioscorea species. The interaction of DIO with bovine serum albumin (BSA) was investigated through spectroscopic methods under simulated physiological conditions. The fluorescence quenching data revealed that the binding of DIO to BSA without or with Co2+ or Zn2+ was a static quenching process. The presence of Co2+ or Zn2+ both increased the static quenching constants K SV and the binding affinity for the BSA-DIO system. In the sight of the competitive experiment and the negative values of ΔH 0 and ΔS 0, DIO bound to site I of BSA mainly through the hydrogen bond and Van der Waals' force. In addition, the conformational changes of BSA were studied by Raman spectra, which revealed that the secondary structure of BSA and microenvironment of the aromatic residues were changed by DIO. The Raman spectra analysis indicated that the changes of conformations, disulfide bridges, and the microenvironment of Tyr, Trp residues of BSA induced by DIO with Co2+ or Zn2+ were different from that without Co2+ or Zn2+.

Characterization of intermolecular interaction between cyanidin‐3‐glucoside and bovine serum albumin: Spectroscopic and molecular docking methods

The intermolecular interaction between cyanidin-3-glucoside (Cy-3-G) and bovine serum albumin (BSA) was investigated using fluorescence, circular dichroism and molecular docking methods. The experimental results revealed that the fluorescence quenching of BSA at 338 nm by Cy-3-G resulted from the formation of Cy-3-G-BSA complex. The number of binding sites (n) for Cy-3-G binding on BSA was approximately equal to 1. The experimental and molecular docking results revealed that after binding Cy-3-G to BSA, Cy-3-G is closer to the Tyr residue than the Trp residue, the secondary structure of BSA almost not change, the binding process of Cy-3-G with BSA is spontaneous, and Cy-3-G can be inserted into the hydrophobic cavity of BSA (site II') in the binding process of Cy-3-G with BSA. Moreover, based on the sign and magnitude of the enthalpy and entropy changes (ΔH(0) = - 29.64 kcal/mol and ΔS(0) = - 69.51 cal/mol K) and the molecular docking results, it can be suggested that the main interaction forces of Cy-3-G with BSA are Van der Waals and hydrogen bonding interactions.

Spectroscopic studies on the interaction between chalcone and bovine serum albumin

The interaction between chalcone and bovine serum albumin (BSA) has been studied by spectroscopic techniques under physiological condition. By the analysis of fluorescence spectrum and fluorescence intensity, it was observed that the chalcone has a strong ability to quench the intrinsic fluorescence with BSA through a static quenching procedure and non-radiation energy transfer were the main reasons for the fluorescence quenching. The association constants of chalcone with BSA were determined at different temperatures based on fluorescence quenching results. The positive entropy change and enthalpy change indicated that the interaction of chalcone and BSA was driven mainly by hydrophobic forces. The process of binding was a spontaneous process in which Gibbs free energy change was negative. The distance, r, between donor (BSA) and acceptor (chalcone) was obtained according to the Forster's theory of non-radiation energy transfer. The UV–vis, CD, FT-IR, synchronous and 3-D spectral results revealed the changes in the secondary structure of BSA upon interaction with chalcone. The effects of some common metal ions on binding of BSA–chalcone complex were also investigated.

Tricalcium phosphate and tricalcium phosphate/polycaprolactone particulate composite for controlled release of protein

β-Tricalcium phosphate (β-TCP) with three different particle size ranges was used to study the effects of particle size and surface area on protein adsorption and release. Polycaprolactone (PCL) coating was applied on the particle systems to investigate its effect on particulate system properties from both structural and application aspects. The maximum loading of 27mg/g was achieved for 100nm particles. Bovine serum albumin (BSA) loading amount was controlled by varying the BSA loading solution concentration, as well as the sample powder's surface area. Increasing the surface area of the delivery powder significantly increased loading and release yield. Unlike the samples with low surface area, the lowest particle size samples showed sigmoidal release profile. This indicated that release was governed by different mechanisms for particles with different sizes. While the majority of samples showed no more than 50% release, the 550nm particles demonstrated 100% release. PCL coating showed no significant ability to attenuate burst release in PBS. However, it led to a steadier release profile as compared to the bare TCP particles. FTIR analysis also proved that the secondary structure of BSA did not change significantly during the adsorption; however, minor denaturation was found during the release. The same results were found when PCL coating was applied on the TCP particles. We envision potential use of TCP and TCP+PCL systems in bone growth factor or orthopedic drug delivery applications in future bone tissue engineering application.

Interactions between U(VI) and bovine serum albumin

Interest in bio-toxicology of uranium resulting from its radioactive heavy metal property has been growing enormously in recent years. The interactions between uranium(VI) [U(VI)] and bovine serum albumin (BSA) at physiological pH were studied by spectroscopic methods. Fuorescence results revealed the formation of BSA–U(VI) complex, the binding constants as well as the number of binding sites were determined. In particular, the effects of U(VI) binding on the secondary structures of BSA were examined by means of Fourier transformation infrared spectroscopy equipped with attenuated total reflection (FT-IR/ATR). It was found that the α-helix component of BSA decreased gradually with increasing concentration of U(VI). In contrast, the β-sheets, turns, and random coil structures all increased correspondingly. Our work would shed light on the possible interaction mechanism between U(VI) and proteins in aqueous solutions.

Assessment of the interaction between fraxinellone and bovine serum albumin by optical spectroscopy and molecular modeling methods

The interaction of fraxinellone with bovine serum albumin (BSA) was investigated by fluorescence spectroscopy, ultraviolet–visible (UV–vis) absorbance, Fourier transform infrared (FT-IR) spectroscopy, and protein-ligand docking studies. The results suggested that the quenching of BSA fluorescence by fraxinellone was a static process. The binding constant Ka at 298K was calculated to be 6.61×104Lmol−1, and the number of binding sites n were given. The thermodynamic parameters, enthalpy change (ΔH) and entropy change (ΔS), at different temperatures were −66.22kJmol−1 and −129.77Jmol−1K−1, respectively. Fraxinellone was bound to BSA in site I, and the distance between fraxinellone and BSA was 2.31nm, as calculated based on the Förster theory of non-radiation energy transfer. Moreover, the obtained UV–vis, synchronous fluorescence, and FT-IR spectra demonstrated that the BSA microenvironment and secondary structure were modified in the presence of fraxinellone. The molecular modeling results are consistent with the displacement and binding mode studies conducted.