Binding Sites Of Human Serum Albumin Research Articles

Binding of human serum albumin with uranyl ion at various pH: an all atom molecular dynamics study

Uranium is routinely handled in various stages of nuclear fuel cycle and its association with human serum albumin (HSA) has been reported in literature, however, their binding characteristics still remains obscure. The present study aims to understand interaction of uranium with HSA by employing all atom molecular dynamics simulation of the HSA-metal ion complex. His67, His247 and Asp249 residues constitute the major binding site of HSA, which capture the uranyl ion (UO2 2+). A total of six sets of initial coordinates are used for Zn2+-HSA and UO2 2+-HSA system at pH = 4, 7.4 and 9, respectively. Enhance sampling method, namely, well-tempered meta-dynamics (WT-MtD) is employed to study the binding and un-binding processes of UO2 2+ and Zn2+ ions. Potential of mean force (PMF) profiles are generated for all the six sets of complexes from the converged WT-MtD run. Various basins and barriers are observed along the (un)binding pathways. Hydrogen bond dynamics and short-range Coulomb interactions are evaluated from the equilibrium run at each basins and barriers for both the ions at all pH values. The binding of UO2 2+ ion with HSA is the result of the dynamical balance between UO2 2+-HSA and UO2 2+-water short range Coulomb interactions. Zn2+ ion interact more strongly than UO2 2+ at all pH through short range Coulomb interactions. PMF values further concludes that UO2 2+ cannot associate to the Zn2+ bound HSA protein but can be captured by free HSA at all pH values i.e. endosomal, alkaline and physiological pH. Communicated by Ramaswamy H. Sarma

Biophysical Characterization of the Interaction between a Transport Human Plasma Protein and the 5,10,15,20-Tetra(pyridine-4-yl)porphyrin.

The interaction between human serum albumin (HSA) and the non-charged synthetic photosensitizer 5,10,15,20-tetra(pyridine-4-yl)porphyrin (4-TPyP) was evaluated by in vitro assays under physiological conditions using spectroscopic techniques (UV-vis, circular dichroism, steady-state, time-resolved, synchronous, and 3D-fluorescence) combined with in silico calculations by molecular docking. The UV-vis and steady-state fluorescence parameters indicated a ground-state association between HSA and 4-TPyP and the absence of any dynamic fluorescence quenching was confirmed by the same average fluorescence lifetime for HSA without (4.76 ± 0.11 ns) and with 4-TPyP (4.79 ± 0.14 ns). Therefore, the Stern–Volmer quenching (KSV) constant reflects the binding affinity, indicating a moderate interaction (104 M−1) being spontaneous (ΔG°= -25.0 kJ/mol at 296 K), enthalpically (ΔH° = -9.31 ± 1.34 kJ/mol), and entropically (ΔS° = 52.9 ± 4.4 J/molK) driven. Binding causes only a very weak perturbation on the secondary structure of albumin. There is just one main binding site in HSA for 4-TPyP (n ≈ 1.0), probably into the subdomain IIA (site I), where the Trp-214 residue can be found. The microenvironment around this fluorophore seems not to be perturbed even with 4-TPyP interacting via hydrogen bonding and van der Waals forces with the amino acid residues in the subdomain IIA.

The aqueous stability and interactions of organoruthenium compounds with serum proteins, cell culture medium, and human serum.

Metal complexes bind to a wide variety of biomolecules and the control of the reactivity is essential when designing anticancer metallodrugs with a specific mode of action in mind. In this study, we used the highly cytotoxic compound [RuII(cym)(8-HQ)Cl] (cym = η6-p-cymene, 8-HQ = 8-hydroxyquinoline), the more inert derivative [RuII(cym)(8-HQ)(PTA)](SO3CF3) (PTA = 1,3,5-triaza-7-phosphaadamantane), and [RuII(cym)(PCA)Cl]Cl (PCA = pyridinecarbothioamide) as a complex with a different coordination environment about the Ru center and investigated their stability, interactions with proteins, and behavior in medium (αMEM) and human serum by capillary zone electrophoresis. The developed method was found to be robust and provides a quick and low-cost technique to monitor the interactions of such complexes with biomolecules. Each complex was found to behave very differently, emphasizing the importance of the choice of ligands and demonstrating the applicability of the developed method. Additionally, the human serum albumin binding site preference of [RuII(cym)(8-HQ)Cl] was investigated through displacement studies, revealing that the compound was able to bind to both sites I and site II, and the type of adducts formed with transferrin was determined by mass spectrometry.

Spectroscopic and Molecular Docking Investigation on the Interaction of Cumin Components with Plasma Protein: Assessment of the Comparative Interactions of Aldehyde and Alcohol with Human Serum Albumin.

The interaction of the important plasma protein, human serum albumin (HSA), with two monoterpenes found in cumin oil, i.e., cuminaldehyde (4-isopropylbenzaldehyde) and cuminol (4-isopropylbenzyl alcohol), was studied in this paper. Both experimental and computational methods were utilized to understand the mechanism of binding. The UV absorption profile of HSA changes in the presence of both cuminaldehyde and cuminol, due to the interaction between HSA with both monoterpenes. The intrinsic fluorescence intensity of HSA was also quenched on the sequential addition of both ligands, due to change in the microenvironment of the fluorophore present in the former. Quenching of HSA by cuminaldehyde was much higher in comparison to that in the presence of cuminol. Fluorescence quenching data were analyzed using modified Stern-Volmer and Lineweaver-Burk methods, which suggested that the binding mechanism was of a static type for both ligands. In both cases, the binding was favored by the domination of hydrophobic as well as hydrogen bonding/Van der Waals forces. Both ligands partially unfolded the secondary structure of HSA, although the effect of cuminaldehyde was more pronounced, as compared to cuminol. The preferred binding site of cuminaldehyde and cuminol inside HSA was also the same; namely, drug binding site 1, located in subdomain IIA. The study showed that cuminaldehyde binds strongly with albumin as compared to its alcohol counterpart, which is due to the more hydrophobic nature of the former.

Interaction of esomeprazole with insulin detemir and human albumin: A potential cause of hypoglycemia

Insulin detemir (IDt) is long-acting insulin whose protraction mechanism is based on a covalently attached fatty acid to an insulin molecule. Utilizing the high affinity of fatty acids towards human serum albumin (HA), the modified detemir molecule binds with good affinity to HA, which functions as a reservoir that leads to a slow and prolonged release of insulin. However, questions were raised over potential interactions between other drugs and IDt through competitive binding on the binding site(s) of HA. In a previous study, concomitant use of esomeprazole (Esom) and erythromycin resulted in severe hypoglycemia, and thus: the drugs including Esom were suggested as enhancers of IDt action through displacing it from its binding site on HA. To further study this possibility, studies utilizing different techniques including, semipermeable membrane dialysis, capillary electrophoresis, UV,NMR spectroscopy, and molecular docking were carried out. Results from various techniques supported the simultaneous binding of Esom along with IDt to HA (i.e., binding in two different sites without signs of competition between the two). Moreover, capillary electrophoresis suggested an increase in the binding affinity of Esom to HA in the presence of IDt (1.9 × 103 Vs 2.7 × 104M−1). Perhaps most interesting was the observation that Esom could bind directly to IDt which was evidenced by all the employed techniques. Direct binding of Esom to IDt, might explain the enhancement in insulin action associated with the concomitant use of Esom. Therefore, Esom might represent a leading insulin-sensitizing compound that might lead to more effective insulin enhancing and less unwanted effects.

Investigation of Pazopanib and Human Serum Albumin Interaction Using Spectroscopic and Molecular Docking Approaches

Pazopanib (PAZ), a tyrosine kinase inhibitor, is used to treat advanced renal cell carcinoma (RCC) and advanced soft tissue sarcoma (STS). The FDA approved PAZ for RCC in 2009 and for STS in 2012. The antitumor activity of pazopanib, according to the degree of inhibition, shows different results depending on the dose. Renal cell carcinoma is the most sensitive carcinoma to pazopanib, with 77% inhibition at the 10 mg/kg dose. Clinical studies have shown 53% to 65% inhibition in carcinomas such as breast carcinoma, prostate carcinoma, and melanoma. Plasma proteins such as human serum albumin (HSA) have a critical role in transporting and storing bioactive components. This feature of HSA is very important for the development of cancer therapy. Here, we investigated the interaction between PAZ and HSA to evaluate their binding strength, binding types, and conformational change in HSA. We used spectroscopic methods to assess the drug–protein interaction. Fluorescence measurements revealed that the interaction of PAZ with HSA occurred via the static quenching mechanism. The calculated binding number and binding constants were 1.041 and 1.436 × 106 M−1, respectively, at 298.15 K based on fluorescence screening. The high binding constant and calculated Gibbs free energy at different temperatures showed spontaneous and strong binding. Circular dichroism measurements showed that the α-helix structure of HSA was retained as the secondary structure, with a slight reduction in its percentage after adding PAZ. Furthermore, molecular modeling studies suggested that the docking score of PAZ is higher than those of bicalutamide and ibuprofen, the drugs that were chosen as model competitors against PAZ. Accordingly, PAZ was found to replace bicalutamide and ibuprofen on the HSA binding site, which was also confirmed by UV absorption spectroscopy.

Glycation Alters the Fatty Acid Binding Capacity of Human Serum Albumin.

Glycation significantly alters the physicochemical and biofunctional properties of proteins in foods and in vivo. In the present study, human serum albumin (HSA) as the major transporter of fatty acids was modified with glyoxal under physiological conditions. Reversibly albumin-bound glyoxal was removed, and advanced glycation end products were quantitated by liquid chromatography-tandem mass spectrometry (LC-MS/MS). The total modification of protein-bound lysine and arginine residues reached up to 4.2 and 9.6%, respectively. The impact of these modifications on the transport capacity of long-chain fatty acids was characterized by spin-labeled fatty acid probes via electron paramagnetic resonance spectroscopy. With increasing degree of glycation, the equivalence of the seven binding sites of native HSA with a dissociation constant of 0.74 ± 0.09 μM was set off with only the three high-affinity sites 2, 4, and 5 remaining (0.46 ± 0.07 μM). The other four sites were shifted to low affinities with significantly higher dissociation constants (1.32 ± 0.35 μM). Tryptic peptide mapping enabled us to relate these findings to molecular changes at specific binding sites. Modification hotspots identified were lysine 351, 286, 159 and arginine 144, 485, 117. Further investigation of plasma protein samples of uremic patients vs healthy controls gave first insights into the in vivo situation.

Synthesis, characterization and investigating the binding mechanism of novel coumarin derivatives with human serum albumin: Spectroscopic and computational approach

Two 4-hydroxycoumarin derivatives: (E)-3-(1-((4‑hydroxy-3-methoxyphenyl)amino) -ethylidene)chromane-2,4‑dione (L1) and (E)-3-(1-((3‑hydroxy-4-methoxyphenyl)-amino)ethylidene) chromane-2,4‑dione (L2), were prepared and structurally characterized by spectroscopic techniques in combination with the B3LYP-D3BJ theoretical method. The interactions between newly synthesized compounds and human serum albumin (HSA) were investigated under physiological conditions at 296,303, and 310 K by fluorescence and absorption spectroscopy, molecular docking, and molecular dynamic simulations. The results of absorption and fluorescence spectral analysis showed that ligands quenched HSA fluorescence through a static process. The corresponding thermodynamic parameters ΔH0, ΔS0, and ΔG0 were calculated according to Van't Hoff's equation. The obtained results indicated that compounds bind spontaneously to HSA mainly by van der Waals's forces and through hydrogen bonds. Ligand-competitive displacement experiments, using known site-specific ligands for HSA's binding sites (I and II) suggest that ligands had a higher affinity for site I (subdomain IIA). The results of the computational analysis follow the experimental data, and the obtained results suggest that the investigated compounds show a good binding affinity according to the HSA receptor, which will be useful for future studies related to rational drug design.

Structural arrangement of the VH and VL domains in the COBRA™ T-cell engaging single-chain diabody.

COBRA™ (COnditional Bispecific Redirected Activation) T-cell engagers are designed to target solid tumors as a single polypeptide chain prodrug that becomes activated by proteolysis in the tumor microenvironment. One COBRA molecule comprises seven Ig domains: three single-domain antibodies (sdAbs) recognizing a tumor target or human serum albumin (HSA), and CD3ε-binding variable fragment heavy chain (VH) and variable fragment light chain (VL) and their inactivated counterparts, VHi and VLi. Pairing of VH and VL, and VLi and VHi into single-chain variable fragments (Fv) is prevented by shortened inter-domain linkers. Instead, VH and VL are expected to interact with VLi and VHi, respectively, thus making a diabody whose binding to CD3ε on the T-cells is impaired. We analyzed the structure of an epidermal growth factor receptor (EGFR) COBRA in solution using negative stain electron microscopy (EM) and small-angle X-ray scattering (SAXS). We found that this EGFR COBRA forms stable monomers with a very dynamic interdomain arrangement. At most, only five domains at a time appeared ordered, and only one VH-VL pair was found in the Fv orientation. Nonenzymatic posttranslational modifications suggest that the CDR3 loops in the VL-VHi pair are exposed but are buried in the VH-VLi pair. The MMP9 cleavage rate of the prodrug when bound to recombinant EGFR or HSA is not affected, indicating positioning of the MMP9-cleavable linker away from the EGFR and HSA binding sites. Here, we propose a model for EGFR COBRA where VH and VLi form an Fv, and VL and VHi do not, possibly interacting with other Ig domains. SAXS and MMP9 cleavage analyses suggest that all COBRA molecules tested have a similar structural architecture.

Combined spectroscopic and computational approaches for the recognition of bioactive flavonoid 6-hydroxyflavone by human serum albumin: Effects of non-enzymatic glycation in the binding

Serum albumin's stability and function are influenced by several structural changes. Non-enzymatic glycation (NEG) is one of the mechanisms which impairs and changes the physiological function of serum albumins. Advanced glycation end products (AGEs) are produced in long-term hyperglycamia and are responsible for structural cell and tissue failures associated with a variety of health disorders. The objective of our study is to assess the consequences of NEG in the molecular recognition of bioactive flavonoid 6-hydroxyflavone by human serum albumin (HSA) using various spectroscopic and computational analyses. UV–visible study revealed the possibility of ground-state interaction of 6HF with HSA/glycated HSA (gHSA) and the presence of static quenching mechanism have been supported by the quenching parameters derived from fluorescence quenching studies at three temperatures (290, 300 and 310 K). Due to glycation-related modulations around the binding sites of HSA, 6HF's interaction with gHSA has higher binding constants than that with HSA. The negative ΔG° showed the spontaneous nature of binding. Positive ΔS° and negative ΔH° indicated that hydrogen bonding and hydrophobic interactions are responsible for the binding. The percentage α-helix of HSA raised in the presence of 6HF, while the effect on gHSA is minor. Site probe experiments revealed that 6HF occupies the binding sites in subdomain IIA and IIIA, but it prefers binding to subdomain IIIA of gHSA. The outcomes from the molecular docking and MD simulation also correlates with the experimental results. This study analyses the changes in the molecular recognition of 6HF by HSA after glycation, which may aid in the understanding of comparable drug pharmacodynamics and offer crucial information for future therapeutic development.

Interactions of chlorophyll-derived photosensitizers with human serum albumin are determined by the central metal ion

Two structurally similar derivatives of chlorophyll a, chlorophyllide a (Chlide) and zinc-pheophorbide a (Zn-Pheide), differing only in central metal ion (Mg2+ or Zn2+, respectively) substituting the tetrapyrrole ring, were investigated with regard to their binding to human serum albumin (HSA). Chlide and Zn-Pheide are very promising photosensitizers with potential application in photodynamic therapy, therefore it is desirable to investigate their interactions with serum proteins. The studies included absorption and steady-state fluorescence spectroscopy, as well as molecular docking. It was found that both investigated compounds form complexes with HSA. Experimental data revealed two classes of binding sites for each compound. The affinities (Ka) for the first class were in the range of 105 and 106 M−1 for Chlide and Zn-Pheide, respectively, while the second class was characterized by the affinities of the order of 104 M−1 for both derivatives. Molecular docking simulations together with displacement studies revealed that the primary binding site of the studied compounds is the heme site, localized in the subdomain IB, however the best characterized binding sites of HSA, namely the Sudlow’s sites I and II are also involved. The interactions between the derivatives of chlorophyll and HSA were found to be predominantly hydrophobic and to a lesser extent hydrogen bonding. Our results demonstrate that the centrally bound metal ion determines both the affinity and mode of binding to HSA, which may be a feature differentiating these compounds in terms of their pharmacokinetics. Communicated by Ramaswamy H. Sarma

Lysine reactivity profiling reveals molecular insights into human serum albumin-small-molecule drug interactions.

Human serum albumin (HSA) is one of the most important serum carrier proteins that deliver small-molecule drugs to their specific targets. Clarifying the molecular mechanism of the interaction between natural HSA and drugs in an aqueous solution has been a hot topic in pharmaceutical chemistry, clinical medicine, and biochemistry in recent years, but it is still challenging. In this paper, the details of molecular interactions of HSA with a variety of therapeutic drugs including ibuprofen, indomethacin, phenylbutazone, and warfarin are systematically investigated using a mass spectrometry (MS)-based lysine reactivity profiling (LRP) strategy. The results reaffirm that the major ligand binding sites (including Sites I and II) of HSA are located in subdomains IIA and IIIA, while several potential drug-binding areas at subdomain IIIB and α helix IIB-IIIA are newly characterized. The MS-LRP strategy may have important application prospects in pharmacodynamics, pharmacokinetics, and safety evaluation of small-molecule drugs.

Comparative studies on biophysical interactions between 4-dicyanomethylene-2,6-dimethyl-4H-pyran (DDP) with bovine serum albumin (BSA) and human serum albumin (HSA) via photophysical approaches and molecular docking techniques

Photophysical studies of 4-Dicyanomethylene-2,6-Dimethyl-4H-Pyran (DDP) dye with globular proteins, Human Serum Albumin (HSA) and Bovine Serum Albumin (BSA) were carried out in aqueous solution. An isosbestic point resulted on the addition of serum albumins, which signifies a complex or an equilibrium state of DDP dye with albumin. Addition of BSA to DDP dye results in a fluorescence enhancement accompanied with a significant hypsochromic shift, whereas with that of HSA, a fluorescence quenching with a considerable blue shift resulted. Excited state studies of DDP dye with serum albumins portray that the role of binding sites of dye with albumins vary considerably and the nature of interaction is presumably attributed to combined hydrogen-bonding and hydrophobic interactions. Molecular docking studies of DDP dye with albumins and two other derivatives 4-(Dicyanomethylene)-2-methyl-6-(4-dimethylaminostyryl)-4H-pyran (DCM) dye and 4-(Dicyanomethylene)-2-methyl-6-(4-t-buyl)-4H-pyran (DCT) dyes with BSA and HSA elucidates that the hydrogen-bonding interaction accompanied with several hydrophobic, pi–pi an pi–alkyl interactions coexist between dye and albumins. The binding energy, intermolecular energy and stability of the DDP, DCM and DCT dyes through docking techniques with albumins authenticate that the dye predominantly acts as hydrogen-bonding acceptor site and the protein molecule as the donor. DDP dye prefers to exist in four different binding sites of HSA, whereas, in the case of BSA, the most preferred site is found to be hydrophobic domain (site I). Interestingly, the most preferred site of DCT dye is III A subdomain of HSA, whereas DCM dye is oriented towards I B subdomain. DDP and DCT are smaller in size and reside in the domain preferred for smaller ligands (II A and IIIA) as resulted in several drugs-HSA interaction whereas DCM dye which is categorized as medium to larger ligand based on the extended structure resides in the most favoured site IB. Fluorescence techniques in combination with molecular docking methods elucidate binding characteristics and the domain in which the dye resides in a micro heterogeneous environment is established in this study.

Interactions of isoorientin and its Semi-synthetic analogs with human serum albumin

Isoorientin is a C-glycosyl flavone with a wide range of health beneficial effects and inhibits glycogen synthase kinase 3β (GSK-3β) potentially against Alzheimer’s disease. Its semi-synthetic derivatives have greater potency than isoorientin. The present study was aimed to determine the mechanism of interactions of isoorientin and its derivatives with human serum albumin (HSA) using multi-spectroscopic, microscale thermophoresis (MST) and computational studies. Spectra of steady-state fluorescence, UV–Vis, and time-resolved fluorescence indicated that isoorientin and its derivatives quenched the intrinsic fluorescence of HSA through a static quenching process. Isoorientin and its derivatives had a moderate affinity with HSA (Ka 7.7–14.9 × 104 M−1). The binding process was accompanied by an exothermic phenomenon, ΔG° of HSA-isoorientin and its derivatives systems were calculated as from −29.51 kJ mol−1 to −27.87 kJ mol−1. Displacement experiments with site-specific markers revealed that isoorientin and its derivatives bind to HSA at site II (subdomain IIIA) only. A reduction in the α-helical content of HSA-isoorientin and its derivatives complex was observed, because the conformational changes was structurally perturbed by the hydrophilic groups of the compounds. Further molecular modeling studies confirmed that the binding of isoorientin and its derivatives to the site II via hydrophobic interaction. The MST results confirmed the interactions between HSA and the compounds of interest. The esterase-like assay studies indicated that isoorientin and its derivatives shared the same binding site in HSA, and their induced structural changes of HSA may have been caused by partial unfolding of HSA. This work helps to understand transport, distribution, bioactivity, and design of flavonoid-based GSK-3β inhibitors.

Spectroscopic and molecular docking investigation of the binding of a bioactive mercaptobenzimidazole-functionalized Schiff base to human serum albumin

This study was conducted to examine the binding of a newly synthesized Schiff base derived from 2-hydroxy-1-naphthaldehyde and 5-amino-2-mercaptobenzimidazole with human serum albumin (HSA) employing various biophysical techniques. The thermal-based fluorescence quenching data indicated that static quenching occurred between the ligand and HSA. The fluorescence results expose that ligand quenches the intrinsic fluorescence of HSA through a static quenching procedure. The thermodynamic parameters of the binding interaction, obtained using van't Hoff equation indicated the spontaneity of the reaction. The stability of the HSA–ligand complex resulted from hydrogen bonding and hydrophobic interactions, which afforded a substantial binding affinity between ligand and HSA. UV–Vis and circular dichroism data indicated that ligand binding induced conformational changes in HSA. The energy transfer efficiency determined according to Fӧorster's theory. Absorption, distribution, metabolism, and excretion (ADME) and Lipinski’s drug likeness of the ligand was predicted, revealing that it had auspicious physicochemical properties for oral bioavailability. Furthermore, molecular modeling was also employed to determine the location of the ligand in HSA binding sites, which revealed that the ligand interacted with polar and apolar residues of site I (subdomain IIA) of HSA, predominantly through hydrophobic and hydrogen bonding interactions.

Mechanistic and conformational studies on the interaction of human serum albumin with rhodamine B by NMR, spectroscopic and molecular modeling methods

Rhodamine B (RB), an important congeneric form of rhodamine dyes, is widely used as a colorant and tracing agent, and recent studies have shown that it is potentially harmful to wildlife and human health. To assess the toxicity of RB at the molecular level, the interaction between human serum albumin (HSA) and RB was investigated using NMR, multispectroscopic methods in combination with computational approaches. The data of NMR indicated that RB indeed bound to HSA, in which the benzene rings were spatially close to the binding site of HSA. The fluorescence results showed that RB had a moderate binding affinity to HSA and strongly quenched the intrinsic fluorescence of HSA through a static quenching mechanism. Analysis of thermodynamic parameters and the effect of ionic strength revealed that electrostatic, hydrophobic and hydrogen bond interactions played an important role in the complex formation. And the main binding region of RB was located near site I of HSA. Furthermore, the microenvironment and conformation of HSA were changed in the presence of RB, which was confirmed by the results obtained from CD, FT-IR, and synchronous fluorescence. Finally, the optimal binding mode of RB to HSA and the stability of HSA-RB complex were studied through molecular modeling methods. In short, this work would provide significant insights for understanding the transport, distribution and toxicity effect of RB when it spread in human body.

Characterization of the interaction of diosgenin with human serum albumin and α1-acid glycoprotein using biophysical and bioinformatic tools

Diosgenin (DIO), a steroidal sapogenin with high therapeutic potential, has been shown to exhibit a myriad of pharmacologically significant properties and is commonly used as a template for synthesizing steroidal drugs. The efficacy of bioactive compounds as a potential drug depends mainly upon their interaction with proteins in the circulatory system. Therefore, the interaction of DIO with the major carrier proteins in humans, i.e. human serum albumin (HSA) and α 1 -acid glycoprotein (AAG), was characterized using multispectroscopic and molecular docking techniques. Analysis of fluorescence enhancement data showed intermediate binding affinity for both DIO–HSA and DIO–AAG interactions. Circular dichroism spectral results revealed minimal conformational alterations in HSA upon DIO binding, but the effect on AAG was more significant. Competitive ligand displacement results suggested that DIO does not bind strongly to the main drug binding sites of HSA. However, DIO was found to favor binding to subdomain IB of HSA based on molecular docking data. For AAG, the binding location of DIO was determined for the two variants of AAG (A and F1*S variants) using molecular docking. For variant A, the binding site was situated in lobes I and II of the central binding pocket of the protein, while in variant F1*S, DIO was docked to a cleft on the protein surface. Hydrophobic interactions and van der Waals forces were mainly responsible for complexation between DIO and both proteins. Additionally, there was involvement of hydrogen bonding between DIO and both variants of AAG, but not between DIO and HSA. • AAG binds to DIO with slightly stronger affinity than HSA. • Hydrophobic forces are mainly responsible for DIO binding to both proteins. • DIO binds to HSA at subdomain IB. • DIO binds to variant F1*S of AAG at a cleft at the surface. • DIO binds to variant A of AAG at the central cavity.

How does cholinium cation surpass tetraethylammonium cation in amino acid-based ionic liquids for thermal and structural stability of serum albumins?

In this study, we report how similarly two serum albumins (bovine serum albumin (BSA) and human serum albumin (HSA)) respond in the presence of different concentration of aromatic amino acid based ionic liquids (AAILs), which are cholinium tryptophan [CHO][Trp]IL and tetraethylammonium tryptophan [TEA][Trp]IL. Extended results of thermodynamic stability indicate the extent to which both serum albumins differ in their thermal stability despite having structural similarity in presence of AAILs. To efficiently quantify the results, biomolecular interactions studies were carried out between serum albumins and AAILs with the help of differential scanning calorimetry (DSC), dynamic light scattering (DLS) and various spectroscopic techniques. DSC results illustrated that both AAILs are increasing the thermal stability of BSA and HSA, as per transition temperature (Tm) values, BSA (65.51 to 72.46 °C) and HSA (65.46 to 75.97 °C) have more thermal stability in the presence of [CHO][Trp]IL as compare to [TEA][Trp]IL, BSA (65.51 to 69.75 °C) and HSA (65.46 to 72.08 °C). Secondary structure results obtained using Dichroweb software and selcon calculations. Furthermore, to illustrate the specific binding of AAIL's cations or anions with the binding sites of BSA and HSA, the molecular docking studies were also performed using Molegro trail version v 6.0.

Cycloplatinated(II) Derivatives of Mercaptopurine Capable of Binding Interactions with HSA/DNA.

In this study, two new bis-cyclometalated Pt(II) complexes, [Pt(C^N)(S^N)] [S^N = deprotonated 6-mercaptopurine (6-MP) and C^N = deprotonated 2-phenylpyridine (ppy), 2a; C^N = deprotonated benzo[h]quinoline (bhq), 2b], are synthesized by the reaction of [PtR(SMe2)(C^N)] (R = Me or p-MeC6H4) with 1 equiv of 6-mercaptopurine (6-HMP) at room temperature. The complexes are fully characterized using 1H and 13C NMR spectroscopies, electrospray ionization mass spectrometry, and elemental analysis. Biomolecular interaction of complex 2a with human serum albumin (HSA) is studied by fluorescence, UV-vis, and circular dichroism (CD) spectroscopies. The binding constants (Kb) and number of binding sites (n) are evaluated using the Stern-Volmer equation. The intrinsic fluorescence of protein is quenched by a static quenching mechanism, with a binding constant of Kb ∼ 105 reflecting a high affinity of complex 2a for HSA. The thermodynamic parameters (ΔH°, ΔG°, and ΔS°) indicate that the interaction is a spontaneous process and hydrophobic forces play a main role in the reaction. The displacement experiments demonstrate that the reactive binding sites of HSA to complex 2a are mainly located within its hydrophobic cavity in subdomain IIA (site I). Synchronous fluorescence spectra reveal that complex 2a affected the microenvironment of tryptophan-214 residues in subdomain IIA of HSA. In the case of interaction of complex 2b and HSA, because of overlapping of the emission spectra of complex 2b with HSA, chemometric approaches are applied. The results indicate significant interaction between the tryptophan residue of HSA and complex 2b. Moreover, the binding of Pt(II) complexes 2a and 2b causes a reduction of the α-helix content of HSA, as obtained by far-UV CD spectroscopy. The average binding distance (r) between Pt(II) complexes and HSA is obtained by Förster's resonance energy-transfer theory. Also, a molecular docking simulation reveals that π-π-stacking and hydrophobic interactions between these complexes and HSA are significant. Furthermore, the interactions of platinum complexes, 2, with calf-thymus DNA (CT-DNA) are investigated. The UV-vis results and ethidium bromide competitive studies support an intercalative interaction of both Pt(II) complexes with DNA. The new complexes 2 are also screened for anticancer activities. The results show that complexes 2 exhibit significant anticancer activity against the K562 (chronic myelogenous leukemia) cell line.

The displacement study of 99m Tc-DTPA-Human serum albumin binding in presence of furosemide and metformin by using equilibrium dialysis and molecular docking.

The 99m Tc-DTPA (Technetium99m diethylenetriaminepentaacetic acid), is a radiopharmaceutical used in renal scintigraphy. The human serum albumin (HSA) binding site(s) for the 99m Tc-DTPA have never been characterized. This study will cover in vitro the binding rates of 99m Tc-DTPA on HSA and the 99m Tc-DTPA competition interactions with two drugs having known human serum albumin binding sites. Furosemide (FUR) and metformin (MET) were added to 99m Tc-DTPA solution (weight ratios 1/1 vol:vol) followed by the quantification of 99m Tc-DTPA binding rates to HSA (40 g/L) using equilibrium dialysis and the qualification of this binding using Molecular Modeling methods. The 99m Tc-DTPA binding rates to human serum albumin increased with the highest concentration. Both drugs FUR and MET displaced 99m Tc-DTPA binding. 99m Tc-DTPA could bind to human serum albumin in many locations in site I and I-II, but strongly bound to site I through hydrogen bonds.