Subdomain IIIA Research Articles

Enantioselective effects of chiral profenofos on the conformation for human serum albumin

Profenofos, as a typical chiral organophosphorus pesticide, can cause various environmental problems and even endanger human health when used in excess. The toxicity of chiral profenofos was investigated through multispectral analysis, molecular docking, and density functional theory (DFT), employing human serum albumin (HSA) as the model protein. Fluorescence titration and lifetime measurements demonstrated that the interaction between chiral profenofos and HSA involves static quenching. Chiral profenofos forms a 1:1 complex with HSA at site II (subdomain IIIA), primarily driven by hydrophobic interactions and hydrogen bonds. Notably, the binding efficacy diminishes as temperature increases. Spectroscopic analyses confirm that chiral profenofos alters the microenvironment and structure of HSA, with the R-enantiomer exerting a greater impact than the S-enantiomer. Consequently, the toxicological implications of the R-profenofos is significantly more pronounced. Investigating the molecular-level toxic effects of chiral pesticides enhances the thoroughness of pesticide assessments, aids in understanding their distribution, metabolism, and associated risks, and facilitates the development of mitigation strategies.

Affinity of endosulfan and HCB with human serum albumin affects serum concentrations in a general population

Endosulfan and hexachlorobenzene (HCB) were previously widely used organochlorine pesticides. Much attention has been paid to trends in human exposure to endosulfan and HCB and to the transport and distribution of endosulfan and HCB within the body. We determined endosulfan I, endosulfan II, and HCB concentrations in serum samples collected from the general population of Wuxi in Jiangsu Province (China) between 2012 and 2016. The endosulfan concentrations varied little throughout the study period, but the HCB concentrations increased. The HCB concentrations were significantly higher in serum from overweight people and smokers than in serum from normal weight people (p = 0.028) and non-smokers (p = 0.011), respectively. HCB binds to human serum albumin (HSA) and can form halogen bonds to CYS-461 and ARG-484 at site II (subdomain IIIA). Fatty acids and nicotine increase the affinity between HCB and HSA, explaining the higher HCB concentrations in serum from overweight people and smokers than normal weight people and non-smokers, respectively.

Mechanism understanding of PIKfyve inhibitor YM201636 with human serum albumin: Insights from molecular modeling and multiple spectroscopic techniques.

YM201636 is the potent PIKfyve inhibitor that is being actively investigated for liver cancer efficacy. In this study, computer simulations and experiments were conducted to investigate the interaction mechanism between YM201636 and the transport protein HSA. Results indicated that YM201636 is stably bound between the subdomains IIA and IIIA of HSA, supported by site marker displacement experiments. YM201636 quenched the endogenous fluorescence of HSA by static quenching since a decrease in quenching constants was observed from 7.74 to 2.39 × 104 M-1. UV-vis and time-resolved fluorescence spectroscopy confirmed the YM201636-HSA complex formation and this binding followed a static mechanism. Thermodynamic parameters ΔG, ΔH, and ΔS obtained negative values suggesting the binding was a spontaneous process driven by Van der Waals interactions and hydrogen binding. Binding constants ranged between 5.71 and 0.33 × 104 M-1, which demonstrated a moderately strong affinity of YM201636 to HSA. CD, synchronous, and 3D fluorescence spectroscopy revealed that YM201636 showed a slight change in secondary structure. The increase of Kapp and a decrease of PSH with YM201636 addition showed that YM201636 changed the surface hydrophobicity of HSA. The research provides reasonable models helping us further understand the transportation and distribution of YM201636 when it absorbs into the blood circulatory system.

Revisiting and Updating the Interaction between Human Serum Albumin and the Non-Steroidal Anti-Inflammatory Drugs Ketoprofen and Ketorolac.

Ketoprofen (KTF) and ketorolac (KTL) are among the most primarily used non-steroidal anti-inflammatory drugs (NSAIDs) in humans to alleviate moderate pain and to treat inflammation. Their binding affinity with albumin (the main globular protein responsible for the biodistribution of drugs in the bloodstream) was previously determined by spectroscopy without considering some conventional pitfalls. Thus, the present work updates the biophysical characterization of the interactions of HSA:KTF and HSA:KTL by 1H saturation-transfer difference nuclear magnetic resonance (1H STD-NMR), ultraviolet (UV) absorption, circular dichroism (CD), steady-state, and time-resolved fluorescence spectroscopies combined with in silico calculations. The binding of HSA:NSAIDs is spontaneous, endothermic, and entropically driven, leading to a conformational rearrangement of HSA with a slight decrease in the α-helix content (7.1% to 7.6%). The predominance of the static quenching mechanism (ground-state association) was identified. Thus, both Stern-Volmer quenching constant (KSV) and binding constant (Kb) values enabled the determination of the binding affinity. In this sense, the KSV and Kb values were found in the order of 104 M-1 at human body temperature, indicating moderate binding affinity with differences in the range of 0.7- and 3.4-fold between KTF and KTL, which agree with the previously reported experimental pharmacokinetic profile. According to 1H STD-NMR data combined with in silico calculations, the aromatic groups in relation to the aliphatic moiety of the drugs interact preferentially with HSA into subdomain IIIA (site II) and are stabilized by interactions via hydrogen bonding and hydrophobic forces. In general, the data obtained in this study have been revised and updated in comparison to those previously reported by other authors who did not account for inner filter corrections, spectral backgrounds, or the identification of the primary mathematical approach for determining the binding affinity of HSA:KTF and HSA:KTL.

On surface binding of serum albumin by harmane: An in vitro spectroscopic and theoretical analysis

Harmane, a naturally occurring beta-carboline alkaloid found in plants and animals, has sparked interest due to its potential biological impacts, specifically its neuro-pharmacological properties and its link with neurodegenerative ailments. This work studies the interaction between the bovine serum albumin (BSA) and the Harmane via multi-spectroscopic and computational analyses. Spectroscopic titrations at different temperatures and concentrations of the salt showed that Harmane forms a stable ground-state complex revealed by a static quenching of the fluorescence spectra. Thermodynamic characteristics of the binding show that BSA and Harmane have a 1:1 binding stoichiometry. The negative change in the Gibbs free energy (ΔG) indicates that the interaction is spontaneous, whereas the positive values of the change in entropy (ΔS) and the change in enthalpy (ΔH) indicate that hydrophobic forces predominate in the binding. The protein surface hydrophobicity index confirms significant surface hydrophobicity. The study also demonstrates the impact of metal ions on protein structure. According to molecular docking studies, Harmane binds to BSA in site II, domain III, subdomain IIIA. Molecular Dynamic (MD) modeling was used to reconfirm the perturbation in the stability and structure of the protein due to Harmane binding at different time frames.

Interaction of different chloro-substituted phenylurea herbicides (diuron and chlortoluron) with bovine serum albumin: Insights from multispectral study

In this work, the interaction between different chloro-substituted phenylurea herbicides (diuron (DIU) and chlortoluron (CHL)) and BSA were investigated and compared at three different temperatures (283 K, 298 K and 310 K) adopting UV–vis, fluorescence, and circular dichroism spectra. The quenching mechanism of the interaction was also proposed. The energy transfer between BSA and DIU/CHL was investigated. The binding sites of DIU/CHL and BSA and the variations in the microenvironment of amino acid residues were studied. The changes of the secondary structure of BSA were analyzed. The results indicate that both DIU and CHL can significantly interact with BSA, and the degree of the interaction between DIU/CHL and BSA increases with the increase of the DIU/CHL concentration. The fluorescence quenching of BSA by DIU/CHL results from the combination of static and dynamic quenching. The DIU/CHL has a weak to moderate binding affinity for BSA, and the binding stoichiometry is 1:1. Their binding processes are spontaneous, and hydrophobic interaction, hydrogen bonds and van der Waals forces are the main interaction forces. DIU/CHL has higher affinity for subdomain IIA (Site I) of BSA than subdomain IIIA (Site II), and also interacts with tryptophan more than tyrosine residues. The energy transfer can occur from BSA to DIU/CHL. By comparison, the strength of the interaction of DIU-BSA is always greater than that of CHL-BSA, and DIU can destroy the secondary structure of BSA molecules greater than CHL and thus the potential toxicity of DIU is higher due to DIU with more chlorine substituents than CHL. It is expected that this study on the interaction can offer in-depth insights into the toxicity of phenylurea herbicides, as well as their impact on human and animal health at the molecular level.

New Ru(II)-p-cymene compounds bearing indomethacin and indomethacin-pyridineamide ligands: synthesis, characterization, computational studies and investigation of their interactions with the Human Serum Albumin

The research interest in ruthenium(II) organometallic compounds has continually increased due to their promising performance in biological assays targeting diverse medicinal applications. This work reports the interactions of the [Ru(II)-η6-p-cymene] framework with the indomethacin (HL1) drug and with its modified analogue indomethacin-pyridineamide (L2). Two new half-sandwich organometallics, [Ru(η6-p-cymene)(L1)Cl] (1) and [Ru(η6-p-cymene)(L2)Cl2] (2), were successfully synthesized and fully characterized by ESI-MS, NMR, ATR-FTIR and UV/VIS spectroscopy. The chemical and electronic structures were corroborated by DFT computational calculations. Compound 2 was chemically stable, keeping the L2 ligand in the coordination sphere in coordinating solvents, and was investigated for the interaction with the Human Serum Albumin (HSA) protein by fluorescence quenching studies. This compound acted as a quencher to the intrinsic protein fluorescence, mainly through binding-related quenching (static) mechanism, showing high value for the apparent association constant (Ka). The analysis of the thermodynamic parameters revealed spontaneous process, presenting low temperature dependence, with electrostatic forces playing a key role in the HSA-compound 2 adduct binding. Site competitive studies suggested that the organometallic 2 may compete for the binding site of ibuprofen (subdomain IIIA) and also for the binding site of Ru(III) naked ion (subdomain IB or IIA). Molecular docking simulations showed that both Ru(II)-arenes may interact through hydrogen bonding with arginine residues, and exhibit high affinity for the microenvironment located in subdomains IIIA and IB. These findings suggest that the HSA protein might be a potential carrier for these Ru(II)-organometallic compounds in the blood plasma.

Insights into interaction of triazine herbicides with three kinds of different alkyl groups (simetryne, ametryn and terbutryn) with human serum albumin via multi-spectral analysis

In this study, the interaction of triazine herbicides with three kinds of different alkyl groups (simetryne, ametryn and terbutryn) with human serum albumin (HSA) are investigated through UV–vis, fluorescence, and circular dichroism (CD) spectra. The mechanisms on the fluorescence quenching of HSA initiated by triazine herbicides are obtained using Stern-Volmer, Lineweaver-Burk and Double logarithm equations. The quenching rate constant (Kq), Stern-Volmer quenching constant (Ksv), binding constant (KA), thermodynamic parameters such as enthalpy change (∆H), entropy change (∆S) and Gibbs free energy (∆G) and number of binding site (n) are calculated and compared. The variations in the microenvironment of amino acid residues are studied by synchronous fluorescence spectroscopy. The binding sites and subdomains are identified using warfarin and ibuprofen as site probes. The conformational changes of HSA are measured using CD spectra. The results reveal that the triazine herbicides with different alkyl groups can interact with HSA by static quenching. The combination of the three herbicides and HSA are equally proportional, and the binding processes are spontaneous. Hydrophobic interaction forces play important roles in simetryne-HSA and ametryn-HSA, while the interaction of terbutryn-HSA is Van der Waals forces and hydrogen bonding. Moreover, the three herbicides can bind to HSA at site I (sub-domain IIA) more than site II (subdomain IIIA), and combine with tryptophan (Trp) more easily than tyrosine (Tyr) residues, respectively. By comparison, the order of interaction strength is terbutryn-HSA > ametryn-HSA > simetryne-HSA. Terbutryn can destroy the secondary structure of HSA more than simetryne and ametryn, and the potential toxicity of terbutryn is higher. It is expected that the interactions of triazine herbicides with HSA via multi-spectral analysis can offer some valuable information for studying the toxicity and the harm of triazine herbicides on human health at molecular level in life science.

Insights into the effect of glucose on the binding between human serum albumin and the nonsteroidal anti-inflammatory drug nimesulide

Glucose interacts with human serum albumin (HSA, the main protein responsible for the biodistribution of drugs in the bloodstream) and consequently affects the binding capacity of exogenous compounds. Thus, in this work, the interactive profile between HSA and the anti-inflammatory drug nimesulide (NMD, used mainly by patients with diabetic neuropathy to relieve acute or chronic pains) was characterized in nonglycemic, normoglycemic (80 mg/dL), and hyperglycemic (320 mg/dL) conditions by biophysics techniques. There is a spontaneous and ground-state association HSA:NMD under physiological conditions. Therefore, the Stern-Volmer constant (Ksv) can also be used to estimate the binding affinity. The Ksv values for nonglycemic, normoglycemic, and hyperglycemic conditions are around 104 M−1, indicating a moderate affinity of NMD to albumin that was slightly improved by glucose levels. Additionally, the binding is enthalpically and entropically driven mainly into subdomains IIA or IIIA. The binding perturbs weakly the α-helix content of albumin, however, glucose potentially stabilizes the tertiary structure, decreasing the structural perturbation upon NMD binding and improves the complex HSA:NMD stability. Overall, the biophysical characterization indicated that glucose levels might slightly positively impact the pharmacokinetic profile of NMD, allowing NMD to achieve its therapeutical potential without affecting drastically its effective dosages.

Critical new insights into the interactions of hexafluoropropylene oxide-dimer acid (GenX or HFPO-DA) with albumin at molecular and cellular levels

A key characteristic to be elucidated, to address the harmful health risks of environmental perfluorinated alkyl substances (PFAS), is their binding modes to serum albumin, the most abundant protein in blood. Hexafluoropropylene oxide-dimer acid (GenX or HFPO-DA) is a new industrial replacement for the widespread linear long-chain PFAS. However, the detailed interaction of new-generation short-chain PFAS with albumin is still lacking. Herein, the binding characteristics of bovine serum albumin (BSA) to GenX were explored at the molecular and cellular levels. It was found that this branched short-chain GenX could bind to BSA with affinity lower than that of legacy linear long-chain perfluorooctanoic acid (PFOA). Site marker competitive study and molecular docking simulation revealed that GenX interacted with subdomain IIIA to form BSA-GenX complex. Consistent with its weaker affinity to albumin protein, the cytotoxicity of branched short-chain GenX was less susceptible to BSA binding compared with that of the linear long-chain PFOA. In contrast to the significant effects of strong BSA-PFOA interaction, the weak affinity of BSA-GenX binding did not influence the structure of protein and the cytotoxicity of GenX. The detailed characterization and direct comparisons of serum albumin interaction with new generation short-chain GenX will provide a better understanding for the toxicological properties of this new alternative.

Deciphering the intriguing molecular recognition of a novel indene-phloroglucinol tethered compound with human serum albumin using multi-spectroscopy and molecular docking studies

A new indene-phloroglucinol tethered compound IPC1 was synthesized in our laboratory in three steps. The interaction of the final compound IPC1 with human serum albumin (HSA) was studied using multi-spectroscopic and molecular docking methods. UV–vis and fluorescence studies confirmed the spontaneous formation of ground state complex and the values of the binding constants were found to be in the order of 104 M−1 (4.727×104 M−1 at 290.15 K). Positive ΔS° (+112.797 J K−1 mol−1) and ΔH° (+6.756 KJ mol−1) indicate the involvement of hydrophobic interaction as the major contributing force in the complex formation process. Site marker studies suggested that the compound IPC1 binds near the subdomain IIIA of HSA, which was further supported by fluorescence spectroscopic and molecular docking analysis.

Decoding the intermolecular recognition mode of a potent anticancer drug, abiraterone with human serum albumin: Assessments through spectroscopic and computational techniques

Abiraterone (ABR) is an FDA-approved anticancer drug that possesses repressing properties against the progression of advanced prostate cancer. The combination of bioactive molecules with proteins in blood circulation is the primary determinant of their potential therapeutic effectiveness. Therefore, the interaction characteristics of ABR with the leading carrier protein, human serum albumin (HSA) was evaluated using multi-spectroscopic and computational techniques. The obtained results showed enhancement of HSA fluorescence upon ABR addition through a static process, and affirmed the complexation between ABR and HSA. Whereas a moderate binding strength in ABR–HSA complexation was manifested, the complex was predicted to be stabilized through hydrophobic interactions, van der Waals forces and hydrogen bonding. Inclusion of ABR to HSA appreciably guarded temperature-induced destabilization of the protein, however, it altered the encompassing medium near Trp and Tyr residues of HSA. ABR was detected to favor binding at subdomain IIIA (Sudlow's site II) of HSA and the constancy of the ABR–HSA complex was revealed.

Model study of the protein-ligand binding in the development of hypersensitivity to folic acid and its analogs

Folic acid (FA) plays a vital role in various metabolic processes, including synthesis and repair of DNA, cell division, the production of red blood cells, and fetal development. However, hypersensitivity to FA and its analogs can occur, leading to various symptomatic manifestations, including shortness of breath, skin rashes, itching, hives, swelling, gastrointestinal disturbances, tachycardia, and anaphylaxis. The mechanism of hypersensitivity to FA and its analogs is not well understood. However, it is known that human serum albumin (HSA) serves as a major pharmacokinetic effector for many substances and drugs, including FA and its analogs such as 5-methyltetrahydrofolic acid (5-MTHF), tetrahydrofolic acid (THFA), and methotrexate (MTX). HSA can interact with these compounds, affecting their distribution and metabolism. The study aimed to study the energetic and topological characteristics of the non-covalent complexes formed between HSA and FA and its analogs in order to obtain more complete information about the potential mechanisms involved in hypersensitivity reactions. Molecular docking was applied to search for the most energetically favorable conformations of the protein-ligand complexes and score the geometries based on their lowest binding energy. The 3D structure of HSA (PDB ID: 1AO6) was used as the docking target, which was obtained from the protein database. The structures of the ligands (FA, 5-MTHF, THFA, and MTX) were downloaded from PubChem, an open chemistry database at the National Institutes of Health. The surface area, volume, and depth of the binding pocket were determined using Proteins Plus. The identification of non-covalent interactions between HSA and the ligands was carried out using the PoseView and DoGSiteScorer web tools. It has been demonstrated that hydrophobic interactions and hydrogen bonds predominantly stabilize all the studied HSA-ligand complexes. Molecular docking analysis revealed that HSA binds the ligands within subdomains IB, IIA, and IIIA, with a binding energy of less than –10.0 kcal/mol. Identifying specific binding sites within the new antigen structures (the complex of HSA with the ligands) can be valuable in determining the energetically favorable binding of epitopes from these antigens to the active sites of IgE antibodies or immune cell receptors.

Synthetic dimethoxyxanthones bind similarly to human serum albumin compared with highly oxygenated xanthones

Spectroscopic studies were carried out for the interaction of the bioactive compounds 2,3-dimethoxyxanthone (1) and 3,4-dimethoxyxanthone (2) with human serum albumin (HSA). The UV absorption, steady-state, and time-resolved fluorescence spectroscopy studies showed that 1 and 2 interact with HSA through a ground-state association. The decrease in the Stern-Volmer constant (KSV) values with increasing temperature, and the bimolecular quenching rate constant (kq) values in the order of 1012 M−1s−1 indicated a static fluorescence quenching mechanism, corroborating with time-resolved fluorescence decays. The association constant (Ka) values in the order of 104 M−1 indicated a moderate interaction between these xanthones and HSA. Circular dichroism experiments showed weak perturbation on the secondary structure of albumin after the interaction with 1 and 2. Thermodynamic parameters suggested that the binding is entropically (for HSA:1, ΔS° 0.012 ± 0.003 and for HSA:2 0.008 ± 0.002 J/molK) and enthalpically (for HSA:1, ΔH° -24.0 ± 0.99 and for HSA:2 -25.6 ± 0.67 kJ/mol) controlled, with a Gibbs free energy change (ΔG°) close to -28.0 kJ/mol in both cases. Competitive drug-displacement and molecular docking results showed that 1 and 2 could interact not only with subdomain IIA, where Trp-214 residue can be found, but also with subdomains IIIA and IB. Hydrogen bonding, electrostatic, and hydrophobic interactions were detected as the main binding forces to stabilize the association HSA:1 and HSA:2. Overall, the dimethoxyxanthones have similar binding affinity compared with highly oxygenated xanthones, e.g., mangiferin, gentiacaulein, and norswertianin.

Unraveling the underlying mechanism of interactions between astaxanthin geometrical isomers and bovine serum albumin

The health benefits of astaxanthin (AST) are related to its geometric isomers. Generally, functional activity is realized by the interactions between active substances and transporters. Hereto, bovine serum albumin (BSA), as a model-binding protein and transporter, is able to recognize and transport isomers of active substances through binding with them. However, differences in the binding mechanism of isomers to BSA may affect the functional activities of isomers through the “binding-transport-activity” chain reaction. Thus, this study sought to elucidate the interactions between AST geometrical isomers and BSA using multi-spectroscopy, surface plasmon resonance and molecular docking. The results showed that Z-AST displayed more interacting amino acid residues and lower thermodynamic parameters than all-E-AST. Meanwhile, the order of binding affinity to BSA was 13Z-AST (1.56 × 10−7 M) > 9Z-AST (2.70 × 10−7 M) > all-E-AST (4.01 × 10−7 M), indicating that Z-AST possessed stronger binding ability to BSA. Moreover, AST isomers were located at the junction between subdomains ⅡA and ⅢA of BSA, and showed the same interaction forces (hydrogen bond and van der Waals force) as well as kinetic processes (slow combination, slow dissociation). These interaction parameters provide valuable insights into their pharmacokinetics in vivo, and it was of great significance to explain the potential differences among AST isomers in functional activities.

Study of Amiloride Binding to Human Serum Albumin: Insights from Thermodynamic, Spectroscopic, and Molecular Docking Investigations

This study was undertaken to investigate the interaction between the sodium channel blocker amiloride (AML) and human serum albumin (HSA). A combination of multi-spectroscopic techniques and computational methods were employed to identify the AML binding site on HSA and the forces responsible for the formation of the HSA–AML complex. Our findings revealed that AML specifically binds to Sudlow’s site II, located in subdomain IIIA of HSA, and that the complex formed is stabilized using van der Waals hydrogen-bonding and hydrophobic interactions. FRET analysis showed that the distance between AML and Trp214 was optimal for efficient quenching. UV-Vis spectroscopy and circular dichroism indicated minor changes in the structure of HSA after AML binding, and molecular dynamics simulations (MDS) conducted over 100 ns provided additional evidence of stable HSA–AML-complex formation. This study enhances understanding of the interaction between AML and HSA and the mechanism responsible.

Ascorbic and salicylic acids modulate the binding interactions of an emergency contraceptive pill levonorgestrel to a model transport protein: Insights from spectroscopy and molecular docking analysis

Serum proteins generally help to transport and distribute drug molecules within the body. In this study, the binding characteristics of bovine serum albumin (BSA) with levonorgestrel (LVG), an emergency contraceptive pill, and the influences of ascorbic acid (ASC) and salicylic acid (SAL) on the binding behaviour and protein structure were elucidated using multi-spectroscopic techniques and molecular docking. The results showed that levonorgestrel decreased BSA intrinsic fluorescence via static quenching mechanism. Binding constant (Ka) values for BSA-LVG complexes were 103 to 104 M-1, indicating their high stabilities. Site probing/docking analysis indicated LVG bound between BSA subdomains IIA and IIIA. UV–visible absorption, Fourier Transform-Infrared and 3D fluorescence spectroscopies affirmed LVG-induced changes in BSA structure, especially in α-helix and β-sheet contents. ASC and SAL influenced BSA conformation for LVG binding and reduced the Ka values by 3.37 and 5.43-folds, respectively. LVG altered the microenvironments of tyrosine residues, interacted with Arg-217, Lys-221, Val-292, Glu-443 etc. within the binding domains. The process was spontaneous (ΔG<0), entropy driven (TΔS>ΔH) and involved van der Waals forces and hydrogen bonding. The findings of the study offered details on the binding interaction between BSA and LVG, and also indicated that prior intake of ASC or SAL could suppress the binding affinity of BSA for levonorgestrel.

Theoretical and experimental investigation of novel quinazoline derivatives: synthesis, photophysical, reactive properties, molecular docking and selective HSA biointeraction

Two new quinazoline derivatives (2a and 2b) were successfully synthesized in this work using the condensation technique in excellent yields. Using spectroscopic techniques and elemental analyses, the compounds were completely characterized. Density functional theory (DFT) computations have been used to examine the title compound’s reactive characteristics. Chemical reactivity was predicted using local reactive descriptors and molecule electrostatic potential. Additionally, Time dependent DFT (TD-DFT) simulations were used to examine the impact of solvents on the photophysical characteristics. The affinity of compounds 2a and 2b for human serum albumin (HSA) was further explored using several electronic spectroscopies. Through static mechanisms, both compounds reduce the intrinsic fluorescence of HSA. It is determined that the HSA-2b complex’s binding constant is significantly greater than the HSA-2a complex. The fluorescence spectrum measurements proved that the HSA underwent structural changes after interaction with these compounds. It was demonstrated by site marker competitive displacement studies that compounds 2a and 2b preferred to bind to site I in HSA subdomain IIA. Additionally, synchronised fluorescence spectra were utilized to analyze how HSA’s conformation changed after interacting with various substances. The molecular docking investigations of these compounds with the three critical HSA binding sites, comprising subdomains IIA, IIIA, and IB, further confirmed the experimental findings. The significant contact between the investigated compounds and HSA was supported by the docking simulations. Communicated by Ramaswamy H. Sarma

Molecular docking of human serum albumin with penicillin G determinants

Background: Human serum albumin (HSA) is the main pharmacokinetic effector of many medications, including penicillin G and its metabolites. An urgent problem of practical medicine is immediate hypersensitivity reactions caused by penicillin toxicity (about 8 % compared to other medications), accompanied by skin pathology, anaphylaxis, and fatality. Objectives: The aim of this study is to describe the structures of penicillin G determinants-HSA complexes and to identify favorable binding sites and the amino acid residues involved in the interaction. Material and Methods: The crystal structure of HSA (ID: 1AO6 from Protein Data Bank) (www.rcsb.org) was chosen as a docking target. Molecular docking methods (AutoDock Tools 1.5.7, AutoDock Vina 1.1.2) were used to gain insight into the interaction of HSA with the major (benzyl penicilloyl G, penicillanic acid) and minor (penicillamine, penicilloic acid, penilloic acid) determinants of penicillin G. Visualization of docking results was implemented in PyMol 2.5. The Protein Plus server (https://proteins.plus) was used to evaluate potential binding pockets. The PLIP tool (https://plip-tool.biotec.tu-dresden.de) was used to identify non-covalent interactions between HSA and its ligands. Results: The molecular docking data indicate that the major determinants of penicillin G are involved in the formation of hydrogen bonds with such HSA residues as Trp214, Arg218, His242, and Asn295; for the minor determinants — Asp108, His146, Tyr148, Ser193, Arg197, Gln204. Both types of determinants are located in the hydrophobic cavity of subdomains IIA and IB. Hydrophobic interactions are present mainly between penicillin G determinants and amino acid residues of subdomain IIIA, such as Ala350, Asp451, Tyr452, and Gln459. Conclusion: The study of penicillin G determinants-HSA complexes is important in the pathogenesis of antibiotic allergy. Identification of specific binding sites can be useful for the development and synthesis of new immunogenic antigens (complexes of major and minor determinants of penicillin G with HSA) that can stimulate the immune system and produce specific antibodies to prevent allergic reactions.

Analysis of the binding mechanism for a water-soluble Pd(II) complex containing β-amino alcohols with HSA applying experimental and computational methods

In the study ahead, the binding interactions of the [Pd (HEAC) Cl2] complex with human serum albumin (HSA) protein have been assayed in vitro (pH= 7.40) utilizing computational and experimental procedures. The mentioned complex was synthesized as a water-soluble complex from {2-((2-((2-hydroxyethyl)amino)ethyl)amino) cyclohexanol} ligand = HEAC. The results of electronic absorption and circular dichroism investigations illustrated that the hydrophobicity of the Tryptophan microenvironment in HSA undergoes the changes by binding to the Pd(II) complex without substantial perturbations on the protein secondary structure. The fluorescence emission spectroscopy analysis revealed that with rising temperature, the quenching constant (K sv) in the Stern–Volmer’s relation decreases; so, it can be said that the interaction process is along with a static quenching mechanism. The values of 2.88 × 105 M−1, and 1.26 represent the binding constant (K b) and the number of the binding sites (n), respectively. The Job graph showed the maximum point at χ = 0.5, which means organizing a new set with 1:1 stoichiometry. Thermodynamic profile (ΔH < 0, ΔS < 0, and ΔG < 0) has affirmed that van der Waals forces and hydrogen bonds have a basic function in the Pd(II) complex-albumin bindings. The ligand-competitive displacement studies utilizing warfarin and ibuprofen have represented that Pd(II) complex interacts with albumin by site II (subdomain IIIA). The computational molecular docking theory approved the results of the site-competitive tests; also, it indicated the existence of hydrogen bonds and van der Waals forces in Pd(II) complex-albumin interactions. Communicated by Ramaswamy H. Sarma