Biophysical insights into the binding characteristics of bovine serum albumin with dipyridamole and the influence of molecular interaction with β cyclodextrin

Abstract

The binding characteristic of anti-platelet drug dipyridamole has been investigated with a transport protein, serum albumin. A multi-spectroscopic approach has been employed, and the results were well supported by in silico molecular docking and simulation studies. The fluorescence quenching of serum albumin at three different temperatures revealed that the mechanism involved is static and the binding constant of the interaction was found to be of the order of 10⁴ M^?1. The reaction was found to be spontaneous and involved hydrophobic interactions. Synchronous, 3D fluorescence and CD spectroscopy indicated a change in conformation of bovine serum albumin (BSA) on interaction with DP. Using site-selective markers, the binding site of DP was found to be in subdomain IB. Molecular docking studies further corroborated these results. Molecular dynamic (MD) simulations showed lower RMSD values on interaction, suggesting the existence of a stable complex between DP and BSA. Furthermore, since β-Cyclodextrin (βCD) is used to improve the solubility of DP in ophthalmic solutions, therefore, the effect of (βCD) on the interaction of BSA and DP was also studied, and it was found that in the presence of βCD, the binding constant for BSA-DP interaction decreased. The present study is an attempt to characterize the transport of DP and to improve its bioavailability, consequently helping in dosage design to achieve optimum therapeutic levels.

Communicated by Ramaswamy H. Sarma     

Elucidating the molecular interaction of serum albumin with nizatidine and the role of β-cyclodextrin: multi-spectroscopic and computational approach

Abstract

Nizatidine is a histamine H2 receptor antagonist which act by inhibiting the production of stomach acid, thereby, finds its application in treating various diseases related to the gastrointestinal tract. Studying albumin–drug interaction is important for understanding the pharmacokinetics and pharmacodynamics of therapeutic candidates. In the present work, the interaction of nizatidine with BSA was investigated by employing multi-spectroscopic and computational studies. The formation of BSA–nizatidine complex was characterised by UV-visible and fluorescence based-spectroscopic studies. Steady-state fluorescence demonstrated the static mode of quenching of BSA by nizatidine. The interaction was spontaneous and nizatidine binds to BSA with a stoichiometry of 1:1. Forster resonance energy transfer calculations revealed that there was a high possibility of energy transfer between nizatidine and BSA. The resultant secondary structural change in BSA on the addition of nizatidine was studied by circular dichroism spectroscopy. Moreover, synchronous and three-dimensional fluorescence spectroscopy was used to determine the conformational changes occurred in the structure of albumin on the binding of nizatidine. Competitive-site marker experiments suggested that nizatidine binds in the Sudlow site II of BSA. Additionally, the effect of β-cyclodextrin as an inclusion compound on the interaction was studied. Furthermore, molecular modelling and simulation studies were performed to corroborate the results obtained above.

Communicated by Ramaswamy H. Sarma     

Inclusion complexation between baicalein and β-cyclodextrin and the influence of β-cyclodextrin on the binding of baicalein with DNA: a spectroscopic approach

This work deals with the commonly studied cyclic oligosaccharide and gains importance as it is entered on a drug delivering carbohydrate and provides insight into the oligosaccharide complex–biomolecular interaction. The binding of a flavone, baicalein, to β-cyclodextrin and calf thymus DNA is studied. The binding of baicalein to calf thymus DNA in the presence of β-cyclodextrin is analysed using the UV–vis absorption and fluorescence spectroscopy. The mode of binding and structure of the baicalein–β-cyclodextrin complex are reported. The role of the structure and the stoichiometry of the inclusion complex of baicalein–β-cyclodextrin in its influence on DNA binding are analysed.

Highlights

? This paper deals with the binding of a flavone, baicalein to β-cyclodextrin and/or DNA.

? The inclusion complexation between baicalein and β-cyclodextrin is analysed.

? The stoichiometry and the binding strength of the inclusion complex is reported.

? The role of β-cyclodextrin in tuning the binding of baicalein to DNA is emphasized.

? Spectroscopic and docking analysis are used to articulate the results.     

Nanopore sensing of γ-cyclodextrin induced host-guest interaction to reverse the binding of perfluorooctanoic acid to human serum albumin

Perfluorooctanoic acid (PFOA) has been one of the most common perfluorochemicals, which are globally pervasive contaminants that are persistent, bioaccumulative, toxic, and have adverse impacts on human health. The highest concentration of PFOA occurs in the blood, where it strongly binds to human serum albumins (HSA). Thus, a method to reverse the HSA-PFOA binding is critical to help facilitate the faster elimination of PFOA from the body to minimize its toxicological effects. Inspired by the remediation effect of cyclodextrin (CD) to PFOA through host-guest interactions, herein, by elucidating inter-molecular interactions using a nanopore sensor, we demonstrated in vitro reversal of the binding of PFOA to HSA using γ-cyclodextrin (γ-CD). The competition behavior for the complexation of PFOA between HSA and γ-CD was discussed in combination with in situ nanopore current recording and nuclear magnetic resonance (NMR) characterization. The present work not only demonstrates the potential therapeutic application of γ-CD for PFOA removal from human blood, but also provides an emerging method for investigating interactions between organic compounds and proteins.     

Binding of acetylated low density lipoprotein and maleylated bovine serum albumin to the rat liver: one or two receptors?

The liver is the major organ involved in clearance of acetylated low density lipoprotein (acetyl-LDL) and maleylated serum albumin (Mal-BSA). Quantitative analysis of the hepatic uptake by sequential scintigraphy in rats shows that the hepatic uptake capacity for Mal-BSA is at least 15 times larger than for acetyl-LDL particles. A membrane-associated M approximately 250,000 daltons hepatic receptor for acetyl-LDL and Mal-BSA was 1450-fold purified from total membrane by Triton X-114 solubilization, chromatography on polyethylenimine cellulose and gel filtration. This receptor incorporated into liposomes displayed a saturable binding of [131I]Mal-BSA with a dissociation constant Kd = 15 nM and to [131I]acetyl-LDL with a dissociation constant Kd = 0.9 nM. The binding of both ligands was sensitive to poly(vinyl sulfate). The purified scavenger receptor system has a binding capacity for [131I]Mal-BSA 20 times larger than for [131I]acetyl-LDL. This is similar to the maximal removal capacity of the rat liver for both ligands in vivo. Binding studies with Mal-BSA, acetyl-LDL and anti-idiotypic receptor antibodies as competitors for [131I]Mal-BSA and [131I]acetyl-LDL binding demonstrate that [131I]Mal-BSA and [131I]acetyl-LDL compete for a common binding site. However, not all of the Mal-BSA binding sites are capable of interacting with acetyl-LDL.     

Binding forces between a novel Schiff base palladium(II) complex and two carrier proteins: human serum albumi and β-lactoglobulin

Ligand binding studies on carrier proteins are crucial in determining the pharmacological properties of drug candidates. Here, a new palladium(II) complex was synthesized and characterized. The in vitro binding studies of this complex with two carrier proteins, human serum albumin (HSA), and β-lactoglobulin (βLG) were investigated by employing biophysical techniques as well as computational modeling. The experimental results showed that the Pd(II) complex interacted with two carrier proteins with moderate binding affinity (K_b ≈ .5 × 10⁴ M^?1 for HSA and .2 × 10³ M^?1 for βLG). Binding of Pd(II) complex to HSA and βLG caused strong fluorescence quenching of both proteins through static quenching mechanism. In two studied systems hydrogen bonds and van der Waals forces were the major stabilizing forces in the drug-protein complex formation. UV–Visible and FT-IR measurements indicated that the binding of above complex to HSA and βLG may induce conformational and micro-environmental changes of two proteins. Protein–ligand docking analysis confirmed that the Pd(II) complex binds to residues located in the subdomain IIA of HSA and site A of βLG. All these experimental and computational results suggest that βLG and HSA might act as carrier protein for Pd(II) complex to deliver it to the target molecules.     

Ligand binding strategies of human serum albumin: How can the cargo be utilized?

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 functional 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, 2010. © 2009 Wiley‐Liss, Inc.     

A comparison of the crosslinking abilities of glutaraldehyde,formaldehyde and α-hydroxyadipaldehyde with bovine serum albumin and casein

Summary The crosslinking effects of formaldehyde, -hydroxyadipaldehyde and glutaraldehyde have been compared by various techniques. Using a micro-Ouchterlony technique with an aldehyde treated bovine serum albumin-rabbit anti-bovine serum albumin system it was found that glutaraldehyde prevented precipitin line formation except at very high titres of antibody. The effects of formaldehyde and -hydroxyadipaldehyde were less marked. Cellulose acetate electrophoresis of aldehyde treated bovine serum albumin showed an increase in mobility compared with the untreated protein. Starch gel electrophoresis of aldehyde fixed liver slices showed no protein loss after glutaraldehyde fixation whereas the other aldehydes permitted proteins to be extracted. Polyacrylamide gel electrophoresis of the aldehyde treated bovine serum albumin showed a little change in mobility after formaldehyde and -hydroxyadipaldehyde treatment and a little polymer formation. Glutaraldehyde on the other hand produced much polymer. These findings were confirmed by gel filtration with Sephadex G-200. Intermolecular crosslinking with glutaraldehyde was dependant on the aldehyde concentration.     

Specific binding of a β-cyclodextrin dimer to the amyloid β peptide modulates the peptide aggregation process

Alzheimer's disease involves progressive neuronal loss. Linked to the disease is the amyloid β (Aβ) peptide, a 38-43-amino acid peptide found in extracellular amyloid plaques in the brain. Cyclodextrins are nontoxic, cone-shaped oligosaccharides with a hydrophilic exterior and a hydrophobic cavity making them suitable hosts for aromatic guest molecules in water. β-Cyclodextrin consists of seven α-d-glucopyranoside units and has been shown to reduce the level of fibrillation and neurotoxicity of Aβ. We have studied the interaction between Aβ and a β-cyclodextrin dimer, consisting of two β-cyclodextrin monomers connected by a flexible linker. The β-cyclodextrin monomer has been found to interact with Aβ(1-40) at sites Y10, F19, and/or F20 with a dissociation constant (K(D)) of 3.9 ± 2.0 mM. Here (1)H-(15)N and (1)H-(13)C heteronuclear single-quantum correlation nuclear magnetic resonance (NMR) spectra show that in addition, the β-cyclodextrin monomer and dimer bind to the histidines. NMR translational diffusion experiments reveal the increased affinity of the β-cyclodextrin dimer (apparent K(D) of 1.1 ± 0.5 mM) for Aβ(1-40) compared to that of the β-cyclodextrin monomer. Kinetic aggregation experiments based on thioflavin T fluorescence indicate that the dimer at 0.05-5 mM decreases the lag time of Aβ aggregation, while a concentration of 10 mM increases the lag time. The β-cyclodextrin monomer at a high concentration decreases the lag time of the aggregation. We conclude that cyclodextrin monomers and dimers have specific, modulating effects on the Aβ(1-40) aggregation process. Transmission electron microscopy shows that the regular fibrillar aggregates formed by Aβ(1-40) alone are replaced by a major fraction of amorphous aggregates in the presence of the β-cyclodextrin dimer.     

Interaction of a surface-active base with the fraction of membrane-bound Williams’ protons

According to the Williams model, the work of mitochondrial respiratory H⁺ pumps gives rise to a fraction of membrane-bound protons (R-protons) that have excess free energy, which is used in the reaction of ATP synthesis. We have earlier managed to detect such a fraction in mitochondria and mitoplasts and to rigorously show (for mitoplasts) that the non-equilibrium R-proton fraction is localized on the surface of the inner membrane. Here we show that a surface-active compound 2,4,6-trichloro-3-pentadecylphenol anion (TCP-C₁₅) selectively interacts with the R-proton fraction, and describe in detail its influence on mitochondrial respiration under conditions of R-proton generation. We also report endogenous regulation of the R-proton fraction volume, which is performed by the phosphate transport system. The results are discussed in terms of the local coupling model.     

Molecular modeling of zinc and copper binding with Alzheimer’s amyloid β-peptide

Aggregation of the amyloid β-peptide (Aβ) into insoluble fibrils is a key pathological event in Alzheimer’s disease. Cu(II) and Zn(II) ions were reported to be able to induce Aβ aggregation at nearly physiological concentrations in vitro. In this study, the binding modes of Cu(II) and Zn(II) in this process were explored by molecular modeling. In the pre-associated Aβ, Nτ atom of imidazole ring of His14, O atom of carbonyl of main-chain and two O atoms of water occupied the four ligand positions of the complex. While in the aggregated form of Aβ, the His13(N)–Metals–His14(N) bridges were formed through metal cross-linking action. These results would be helpful to put insight on revealing the formation mechanism of pathogenic Aβ aggregates in brain.     

The preparation and properties of a highly specific antiserum elicited with 3-dehydrocholylglycine 3-succinyl—bovine serum albumin conjugate

This report describes the synthesis of a new cholylglycine derivative-bovine serum albumin conjugate. The hapten is linked to the carrier protein at the C-3 position, through a hemisuccinate bridge. Antiserum elicited by this antigen is highly specific to cholylglycine. Cross-reactions with free cholic acid (less than 0.1%) or cholyltaurine (0.5%) are minimal.     

Interaction of meropenem with ‘N’ and ‘B’ isoforms of human serum albumin: a spectroscopic and molecular docking study

Carbapenems are used to control the outbreak of β-lactamases expressing bacteria. The effectiveness of drugs is influenced by its interaction with human serum albumin (HSA). Strong binding of carbapenems to HSA may lead to decreased bioavailability of the drug. The non-optimal drug dosage will provide a positive selection pressure on bacteria to develop resistance. Here, we investigated the interaction between meropenem and HSA at physiological pH 7.5 (N-isoform HSA) and non-physiological pH 9.2 (B-isoform HSA). Results showed that meropenem quenches the fluorescence of both ‘N’ and ‘B’ isoforms of HSA (ΔG < 0 and binding constant ~10⁴ M^?1). Electrostatic interactions and van der Waal interactions along with H-bonds stabilized the complex of meropenem with ‘N’ and ‘B’ isoforms of HSA, respectively. Molecular docking results revealed that meropenem binds to HSA near Sudlow’s site II (subdomain IIIA) close to Trp-214 with a contribution of a few residues of subdomain IIA. CD spectroscopy showed a change in the conformation of both the isoforms of HSA upon meropenem binding. The catalytic efficiency of HSA (only N-isoform) on p-nitrophenyl acetate was increased primarily due to a decrease in K_m and an increase in k_cat values. This study provides an insight into the molecular basis of interaction between meropenem and HSA.     

Prediction of Alzheimer’s disease with serum lipid levels in Asian individuals: a meta-analysis

Background: The serum lipid profile has become a routine clinical test and used as an important predictor for Alzheimer’s disease (AD), although its predictive value remains undetermined.

Objective: To evaluate the role of serum lipid levels in predicting the risk of AD.

Methods: Meta-analyses were conducted using Comprehensive Meta-analyses (CMA) software to investigate the association between four conventional serum lipid profile parameters and the risk of AD, focused on samples from Asian.

Results: In total, 3423 AD patients and 6127 healthy participants were involved. The results demonstrated that AD patients showed higher LDL-C and TC levels (SMD?=?0.27, 95% CI: 0.04–0.51, p?=?0.02 for LDL-C; SMD?=?0.25, 95% CI: 0.05–0.46, p?=?0.02 for TC) compared with those of healthy controls. People with higher LDL-C and/or TC levels had an increased risk of AD (OR?=?1.64, 95% CI: 1.07–2.51 for LDL-C and OR?=?1.58, 95% CI: 1.10–2.92 for TC).

Conclusions: This study provided evidence that serum LDL-C and TC levels were associated with the risk of AD in Asian individuals. The routine lipid profile may be useful for AD diagnosis, monitoring and treatment.     

Antibodies directed against the thiomannose moiety of a glycoconjugate of 2-imino-2-methoxyethyl 1-thio-α-d-mannopyranoside and bovine serum albumin

Anti-thiomannose antibodies were induced in rabbits immunized with a glycoconjugate of 2-imino-2-methoxyethyl 1-thio-α-d-mannopyranoside (Man-S) and bovine serum albumin (BSA). Also anti-BSA antibodies directed against the BSA moiety of the glycoconjugate were detected in low concentrations in the immune serum. However, antibodies against the combinatorial epitope of the hapten group and the carrier protein were not detected. The anti-thiomannose and the anti-BSA antibodies were isolated in pure forms by affinity chromatography on Sepharose 4B-bearing thiomannosyl-BSA ligands or BSA ligands. The anti-thiomannose antibodies constituted the major fraction of the antibodies, and these antibodies were isolated in pure form for the first time. The specificity of the thiomannose antibodies was established from data of experiments of periodate oxidation, perpropionic acid oxidation, hapten inhibition, and agar diffusion. Isoelectrofocusing showed that the anti-thiomannose antibody preparation consisted of at least six isomeric proteins, all of which exhibited antibody activity against the glycoconjugate of thiomannose and BSA.     

Levodopa influences the regularity of the ankle joint kinematics in individuals with Parkinson’s disease

The aim of the investigation was to explore the influence of levodopa therapy on the regularity of the structural variations present in the lower extremity joints of individuals with Parkinson’s disease (PD). Ten participants with PD walked on a treadmill during the states of “off” and “on” levodopa. Approximate entropy was used to quantify the regularity of the structural variations present in the joint kinematics. Additionally, a pseudo-periodic surrogation analysis was used to evaluate if changes in the regularity of the joint’s movement were associated with a noisy or deterministic motor process. This investigation provided two key findings. The first was that the structural variations present in ankle joint were more regular with levodopa therapy. The second was that changes in the structural variations were related to a deterministic motor process. This indicated that the variations present in the walking patterns of individuals with PD most likely arose from higher-order neural couplings rather than noise in the motor process. Monitoring the regularity of the structural variations present in gait may help improve the management of PD.     

Going with the flow: a brief history of the study of the honeybee’s navigational ‘odometer’

Honeybees navigate to a food source using a sky-based compass to determine their travel direction, and an odometer to register how far they have travelled. The past 20 years have seen a renewed interest in understanding the nature of the odometer. Early work, pioneered by von Frisch and colleagues, hypothesized that travel distance is measured in terms of the energy that is consumed during the journey. More recent studies suggest that visual cues play a role as well. Specifically, bees appear to gauge travel distance by sensing the extent to which the image of the environment moves in the eye during the journey from the hive to the food source. Most of the evidence indicates that travel distance is measured during the outbound journey. Accumulation of odometric errors is restricted by resetting the odometer every time a prominent landmark is passed. When making detours around large obstacles, the odometer registers the total distance of the path that is flown to the destination, and not the “bee-line” distance. Finally, recent studies are revealing that bees can perform odometry in three dimensions.     

Syntheses OF 15α- and 15β-carboxymethyltestosterone bovine serum albumin conjugates: Characteristics of the antisera to testosterone

Syntheses of 15α- and 15β-carboxymethyltestosterone (15α- and 15β-CMT) were investigated in order to prepare testosterone-bovine serum albumin conjugates for radioimmunoassays of testosterone. A mixture of 15α- and 15β-bis (ethoxycarbonyl)methyl-3β-hydroxy-5-androsten-17-one (IIa and IIb) obtained by a reaction of 3β-hydroxy-5,15-androsta-dien-17-one (I) and sodium diethyl malonate was oxidized to afford a mixture of 15α- and 15β-bis(ethoxycarbonyl)methyl-4-androstene-3, 17-dione (Va and Vb). After the separation by silica gel chromatography, each epimer obtained was hydrolyzed by acid, followed by decarboxylation, and selective reduction of the 17-ketone to give 15α- and 15β-CMT. The antisera, generated in rabbits by immunization with the bovine serum albumin (BSA) conjugates of 15α- and 15β-CMT, respectively, exhibited high specificity for testosterone.     

Complexation of the mycotoxin zearalenone with β-cyclodextrin: Study of the interaction and first promising applications

This work reports the study of the interactions between native and substituted β-cyclodextrins and zearalenone and its derivatives α- and β-zearelonol. The data obtained by fluorescence and NMR experiments suggested that zearalenone, α- and β-zearalenol and cyclodextrins give rise to host-guest complexation, with the inclusion of the phenolic moiety inside the cyclodextrin cavity. The high stability of these complexes induces a high fluorescence enhancement upon complexation. These results have been successfully applied to the spectrofluorimetric determination of zearalenone in maize raw samples, without any chromatographic separation. Presented at the 29^th Mykotoxin-Workshop, Fellbach, Germany, May 14–16, 2007 Financial support: Emilia-Romagna region (project SIQUAL)     

Molecular encapsulation of berberine by a modified β-cyclodextrin and binding of host: guest complex to G-quadruplex DNA

Abstract

The capacity to control quadruplex formation, especially in cancer cells, is captivating and entails a reasonable comprehension of the ligand-G-quadruplex binding. Herein, we report an iminopyrenyl-β-cyclodextrin conjugate interacting with duplex and G-quadrulex DNAs. In addition, the host: guest association of the established G-quadruplex binder, berberine, with the β-cyclodextrin derivative is studied employing 2-D ROESY. NMR, UV-visible, and fluorescence spectroscopic techniques are utilized to explore the β-cyclodextrin conjugate's interaction with the quadruplexes. The Binding constants are accounted for the association of the ligands to each of the DNAs viz., calf thymus DNA (duplex), kit22, telo24, and myc22 (quadruplexes). The modulation of the iminopyrenyl-β-cyclodextrin binding to the DNAs are observed when berberine is loaded in the host molecule. A vivid distinction between the interactions of the ligands with duplex and quadruplex structures is inferred. Berberine-loaded iminopyrenyl-β-cyclodextrin shows a higher affinity for binding to kit22.