Studying the interaction between three synthesized heterocyclic sulfonamide compounds with hemoglobin by spectroscopy and molecular modeling techniques

The interaction between synthesized heterocyclic benzene sulfonamide compounds, N-(7-benzyl-56-biphenyl-2m-tolyl-7H-pyrrolo[23-d]pyrimidine–4–yl)-benzene sulfonamide (HBS₁), N-(7-benzyl-56-biphenyl-2-m-tolyl-7H-pyrrolo[23-d] pyrimidine-4-yl)-4-methyl- benzene sulfonamide (HBS₂), and N-(7-benzyl-56-biphenyl-2-m-tolyl-7H-pyrrolo[23-d]pyrimidine-4-yl)-4-chloro-benzene sulfonamide (HBS₃) with Hb was studied by fluorescence quenching, zeta potentional, circular dichroism, and molecular modeling techniques. The fluorescence spectroscopy experiments were performed in order to study the conformational changes, possibly due to a discrete reorganization of Trp residues during binding between HBS derivatives and Hb. The variation of the K_SV value suggested that hydrophobic and electrostatic interactions were the predominant intermolecular forces stabilizing the complex. The K_SV1 ans K_SV2 values of HBS derivatives with Hb are .6 × 10¹³ and 3 × 10¹³ M^?1 for Hb–HBS₁, 1 × 10¹³ and 4 × 10¹³ M^?1 for Hb–HBS₂, .9 × 10¹³, and 6 × 10¹³ M^?1 for Hb–HBS₃, respectively. The molecular distances between Hb and HBS derivatives in binary and ternary systems were estimated according to Förster’s theory of dipole–dipole non-radiation energy transfer. The quantitative analysis data of circular dichroism spectra demonstrated that the binding of the three HBS derivatives to Hb induced conformational changes in Hb. Changes in the zeta potential of the Hb–HBS derivatives complexes demonstrated a hydrophobic adsorption of the anionic ligand onto the surface of Hb as well as both electrostatic and hydrophobic adsorption in the case of the complex. The modeling data thus confirmed the experimental results. This study is expected to provide important insight into the interaction of Hb with three HBS derivatives to use in various toxicological and therapeutic processes.     

Human hemoglobin adsorption onto colloidal cerium oxide nanoparticles: a new model based on zeta potential and spectroscopy measurements

The nanoparticle (NP)-induced conformational changes of protein and NP agglomeration have gained a remarkable interest in medical and biotechnological fields. Herein, the effect of human hemoglobin (Hb) on the colloidal stability of cerium oxide NP (CNP) was investigated by dynamic light scattering (DLS), zeta potential, and TEM analysis. In addition, the effect of CNP on the heme degradation and structural changes of Hb was studied using fluorescence, circular dichroism (CD), and UV–visible (UV–vis) spectroscopic methods. DLS and TEM analysis showed that the presence of Hb can increase the mean diameter of CNP. Zeta potential measurements revealed that CNP demonstrated a higher charge distribution relative to CNP/Hb complex. Besides, fluorescence studies indicated that two fluorescent heme degradation products are revealed during the interaction of CNP with Hb. Near UV-CD spectroscopy also showed that the microenvironmental changes of heme groups occur after interaction of Hb with CNP. The result of thermal behavior of Hb confirmed the structural changes of protein, which referred to decrease in the Hb stability in the presence of CNP. Indeed, the finding related to structural and functional changes of Hb induced by CNP may be crucial to obtain information regarding the side effects of NPs. Finally, this data reveal much insight into the effects of the interaction on protein structural changes and NP agglomeration, and can correlate the zeta potential of NP-protein complexes with the nature of the principle NP-protein interaction.     

Probing the conformational changes and peroxidase-like activity of cytochrome c upon interaction with iron nanoparticles

Herein, the interaction of iron nanoparticle (Fe-NP) with cytochrome c (Cyt c) was investigated, and a range of techniques such as dynamic light scattering (DLS), zeta potential measurements, static and synchronous fluorescence spectroscopy, near and far circular dichroism (CD) spectroscopy, and ultraviolet–visible (UV–vis) spectroscopy were used to analyze the interaction between Cyt c and Fe-NP. DLS and zeta potential measurements showed that the values of hydrodynamic radius and charge distribution of Fe-NP are 83.95 ± 3.7 nm and 4.5 ± .8 mV, respectively. The fluorescence spectroscopy results demonstrated that the binding of Fe-NP with Cyt c is mediated by hydrogen bonds and van der Waals interactions. Also Fe-NP induced conformational changes in Cyt c and reduced the melting temperature value of Cyt c from 79.18 to 71.33°C. CD experiments of interaction between Fe-NP and Cyt c revealed that the secondary structure of Cyt c with the dominant α-helix structures remained unchanged whereas the tertiary structure and heme position of Cyt c are subjected to remarkable changes. Absorption spectroscopy at 695 nm revealed that Fe-NP considerably disrupt the Fe…S(Met80) bond. In addition, the UV–vis experiment showed the peroxidase-like activity of Cyt c upon interaction with Fe-NP. Hence, the data indicate the Fe-NP results in unfolding of Cyt c and subsequent peroxidase-like activity of denatured species. It was concluded that a comprehensive study of the interaction of Fe-NP with biological system is a crucial step for their potential application as intracellular delivery carriers and medicinal agents.     

Determining the binding site and binding affinity of estradiol to human serum albumin and holo-transferrin: fluorescence spectroscopic,isothermal titration calorimetry and molecular modeling approaches

The interactions between estradiol and two carrier proteins, i.e. human serum albumin (HSA) and holo-transferrin (HTF) in aqueous solution at pH = 7.4 were studied by three-dimensional fluorescence emission spectroscopy, isothermal titration calorimetry (ITC), zeta-potential, resonance light-scattering and molecular modeling. Extensive fluorescence quenching was observed throughout the interaction between the drug and both proteins. Moreover, conformational changes were determined by observing the rearrangement of Trp residues during binding of estradiol with HSA and HTF at different concentrations. ITC experiments revealed that, in the presence of estradiol, both van der Waals forces and hydrogen bonding became predominant. In addition, other binding parameters such as enthalpy and entropy changes were determined by the zeta potential method. Molecular modeling suggested that estradiol was situated within sub-domain IB sited in the hydrophobic cluster in Site I, whereas the drug was located in the N-terminal of HTF where it was hydrogen bonded with Ala 670.     

Destructive effect of anticancer oxali-palladium on heme degradation through the generation of endogenous hydrogen peroxide

The interaction between human hemoglobin (Hb) and oxali-palladium was studied using different spectroscopic methods of UV–vis, fluorescence, circular dichroism (CD), and chemiluminescence at two temperatures of 25 and 37°C. The experimental results showed that both dynamic and static quenching is occurred simultaneously when oxali-palladium quenches the fluorescence of Hb. According to the fluorescence quenching method, the binding site number, apparent binding constant, and corresponding thermodynamic parameters were measured at two temperatures. The values of ΔH°, ΔS°, and ΔG° indicate that process of the formation of oxali-palladium–Hb complex is a spontaneous interaction procedure in which electrostatic interaction plays a major role. In addition, UV–vis and CD results showed that the addition of oxali-palladium changes the conformation of Hb. To evaluate the functional changes of Hb via destruction of the heme structure, fluorescence studies were performed. The results demonstrated that two fluorescent heme degradation products are found during the interaction of oxali-palladium with Hb. Also, the amount of hydrogen peroxide produced in the solution of Hb due to the interaction of oxali-palladium with Hb using chemiluminescence method indicated heme degradation in the protein is occurred. Structural and functional changes induced in Hb via heme degradation are considered as side effects of this synthesized anticancer drug.     

The Interaction of BODIPY with bovine serum albumin and its bilirubin complex

Using fluorescence and electronic spectroscopy the interaction of boron-dipyrromethene complex (BODIPY) with bovine serum albumin (BSA) and its bilirubin macromolecular complex (BR·BSA) in aqueous solution was investigated. The interaction of BODIPY is carried out by the static quenching of protein fluorescence and is predominantly hydrophobic and electrostatic in nature. The values of the binding constants were (61.2 ± 2.8) · 10³ and (6.51 ± 0.3) · 10³ M^?1. The interaction of BODIPY with proteins leads to the observed hypso- and bathochromic shift in BODIPY absorption band. Forster resonance energy transfer theory allowed of determing the donor-ligand distance, which were 2.88 and 2.46 nm for BSA and BR·BSA, respectively. Using synchronous fluorescence spectroscopy it was possible to reveal features of BODIPY influence on conformational changes in protein molecules. It was established that BODIPY more effectively interacts with BSA compared to BR·BSA.     

A comparison study of the interaction between β-lactoglobulin and retinol at two different conditions: spectroscopic and molecular modeling approaches

The interaction between bovin β-Lactoglobulin (β-LG) and retinol at two different pH values was investigated by multispectroscopic, zeta potential, molecular modeling, and conductometry measurements. The steady state and polarization fluorescence spectroscopy revealed that complex formation at two different pH values could occur through a remarkable static quenching. According to fluorescence quenching, one set of binding site at pH 2 and two sets of binding sites at pH 7 were introduced for binding of retinol to β-LG that show the enhancement of saturation score of β-LG to retinol in dimmer condition. The polarization fluorescence analysis represented that there is more affinity between β-LG and retinol at pH 7 rather than at pH 2. The effect of retinol on β-LG was studied by UV-visible, circular dichroism (CD), and synchronous fluorescence, which indicated that retinol induced more structural changes on β-LG at pH 7. β-LG–retinol complex formation at two different pH values was recorded via applying resonance light scattering (RLS) and zeta potential. Conductometry and RLS showed two different behaviors of interaction between β-LG and retinol at two different pH values; therefore, dimmer formation played important roles in different behaviors of interaction between β-LG and retinol. The zeta potential was the implied combination of electrostatic and hydrophobic forces which are involved in β-LG–retinol complex at two different pH values, and the hydrophobic interactions play a dominant role in complex formation. Molecular modeling was approved by all experimental results. The acquired results suggested that monomer and dimmer states of β-LG can be induced by retinol with different behaviors.     

Curcumin specifically binds to the human calcium–calmodulin-dependent protein kinase IV: fluorescence and molecular dynamics simulation studies

Calcium–calmodulin-dependent protein kinase IV (CAMK4) plays significant role in the regulation of calcium-dependent gene expression, and thus, it is involved in varieties of cellular functions such as cell signaling and neuronal survival. On the other hand, curcumin, a naturally occurring yellow bioactive component of turmeric possesses wide spectrum of biological actions, and it is widely used to treat atherosclerosis, diabetes, cancer, and inflammation. It also acts as an antioxidant. Here, we studied the interaction of curcumin with human CAMK4 at pH 7.4 using molecular docking, molecular dynamics (MD) simulations, fluorescence binding, and surface plasmon resonance (SPR) methods. We performed MD simulations for both neutral and anionic forms of CAMK4-curcumin complexes for a reasonably long time (150 ns) to see the overall stability of the protein–ligand complex. Molecular docking studies revealed that the curcumin binds in the large hydrophobic cavity of kinase domain of CAMK4 through several hydrophobic and hydrogen-bonded interactions. Additionally, MD simulations studies contributed in understanding the stability of protein–ligand complex system in aqueous solution and conformational changes in the CAMK4 upon binding of curcumin. A significant increase in the fluorescence intensity at 495 nm was observed (λ_exc = 425 nm), suggesting a strong interaction of curcumin to the CAMK4. A high binding affinity (K_D = 3.7 × 10^?8 ± .03 M) of curcumin for the CAMK4 was measured by SPR further indicating curcumin as a potential ligand for the CAMK4. This study will provide insights into designing a new inspired curcumin derivatives as therapeutic agents against many life-threatening diseases.     

Binding of Janus kinase inhibitor tofacitinib with human serum albumin: multi-technique approach

In this report, we have investigated the binding affinity of tofacitinib with human serum albumin (HSA) under simulated physiological conditions by using UV–visible spectroscopy, fluorescence quenching measurements, dynamic light scattering (DLS), differential scanning calorimetry (DSC) and molecular docking methods. The obtained results demonstrate that fluorescence intensity of HSA gets quenched by tofacitinib and quenching occurs in static manner. Binding parameters calculated from modified Stern–Volmer equation shows that the drug binds to HSA with a binding constant in the order of 10⁵. Synchronous fluorescence data deciphered the change in the microenvironment of tryptophan residue in HSA. UV spectroscopy and DLS measurements deciphered complex formation and reduction in hydrodynamic radii of the protein, respectively. Further DSC results show that tofacitinib increases the thermo stability of HSA. Hydrogen bonding and hydrophobic interaction are the main binding forces between HSA and tofacitinib as revealed by docking results.     

Interaction of two flavonols with fat mass and obesity-associated protein investigated by fluorescence quenching and molecular docking

The binding of two flavonols with fat mass and obesity-associated protein (FTO) was studied using fluorescence spectroscopy, Stern-Volmer kinetics, UV-Vis absorption, and molecular docking. The quenching of FTO fluorescence was determined to be static with binding constants on the order of 10⁴ M^?1. The interaction was studied over three temperatures, and the binding was found to be exothermic with a positive change in entropy. Thermodynamic analysis and molecular modeling suggest that hydrophobic interaction and hydrogen bonding interaction are the main binding force in stabilizing the flavonol–FTO complex.     

Spectroscopic Studies of the Supramolecular Interactions Between Uracil and 5-Hydroxy-6-Methyluracil with Bovine Serum Albumin and its Bilirubin Complex

Using fluorescence and absorption spectroscopy the interaction of bovine serum albumin and its bilirubin complex with uracil and 5-hydroxy-6-methyluracil in phosphate buffer at pH 7.4 was investigated. The parameters of forming intermolecular complexes (binding constants, quenching rate constants, the radius of the quenching sphere and etc.) were determined. The interaction between serum albumin and uracils is carried out by the static quenching of protein fluorescence and has predominantly hydrophobic character. Using synchronous fluorescence spectroscopy the influence of uracil and 5-hydroxy-6-methyluracil on the conformational changes of the protein molecule was studied. Uracils effectively binds to bilirubin-albumin complex compared to free protein, which is caused by the interaction with tetrapyrrolic pigment in macromolecular complex. Molecular docking calculations also being presented.     

Probing the intermolecular interactions into serum albumin and anthraquinone systems: a spectroscopic and docking approach

Intermolecular interaction study of human serum albumin (HSA) with two anthraquinones i.e. danthron and quinizarin has been performed through fluorescence, UV-vis and CD spectroscopy along with docking analysis. The titration of drugs into HSA solution brought about the quenching of fluorescence emission by way of complex formation. The binding constants were found to be 1.51 × 10⁴ L mol^?1 and 1.70 × 10⁴ L mol^?1 at λ_exc = 280 nm while at λ_exc = 295 nm, the values of binding constants were 1.81 × 10⁴ L mol^?1 and 1.90 × 10⁴ L mol^?1 which hinted toward binding of both the drugs in the vicinity of subdomain IIA. Different temperature study revealed the presence of static quenching mechanism. Moreover, more effective quenching of the fluorescence emission was observed at λ_exc = 295 nm which also suggested that both the drug molecule bind nearer to Trp-214. Thermodynamic parameters showed that hydrophobic interaction was the major force behind the binding of drugs. The UV-vis spectroscopy testified the formation of complex in both the systems and primary quenching mechanism as static one. The changes in secondary structure and α-helicity in both the systems were observed by circular dichroism spectroscopy. Furthermore, molecular docking analysis predicted the probable binding site of drugs in subdomain IIA of HSA molecule. The types of amino acid residues surrounding the drug molecule advocated that van der Waals forces, hydrophobic forces and electrostatic forces played a vital role in the stabilization of drug-protein complex formed.     

Binding of the alkaloid aristololactam-β-D-glucoside and daunomycin to human hemoglobin: spectroscopy and calorimetry studies

The interaction of the plant alkaloid aristololactam-β-D-glucoside (ADG) and the anticancer agent daunomycin (DAN) with human hemoglobin was studied by different spectroscopic and calorimetric methods. The binding affinity values of ADG and DAN, estimated from spectroscopic experiments, were 3.79 × 10⁴ and 6.68 × 10⁴ M^?1, respectively. From circular dichroism, 3D fluorescence, and FTIR studies it was observed that, DAN induced stronger conformational changes than ADG in the protein. From synchronous fluorescence spectroscopy results, a pronounced shift in the maximum emission wavelength of tyrosine residues was observed in both cases suggesting that the drugs changed the polarity around tyrosine residues with marginal change around the tryptophan residues. The thermodynamics of the binding interaction analyzed using microcalorimetry presented single binding events that were exothermic in nature in both cases. The binding was driven by large positive standard molar entropy changes with small favorable enthalpy contributions. Negative heat capacity changes in both cases are correlated to the involvement of significant hydrophobic forces in the complexation process. The affinity of DAN to Hb was higher than that of ADG.     

Study of the interaction between DNP and DIDS with human hemoglobin as binary and ternary systems: spectroscopic and molecular modeling investigation

The combination of several drugs is necessary, especially during long-term therapy. A competitive binding of the drugs can cause a decrease in the amount of drugs actually bound to the protein and increase the biologically active fraction of the drug. Here, the interaction between 4,4′-Diisothiocyano-2,2′-stilbenedisulfonic acid (DIDS) and 2,4-Dinitrophenol (DNP) with Hemoglobin (Hb) was investigated by different spectroscopic and molecular modeling techniques. Fluorescence analysis was used to estimate the effect of the DIDS and DNP on Hb as well as to define the binding properties of binary and ternary complexes. The distance r between donor and acceptor was obtained by the FRET and found to be 2.25 and 2.13 nm for DIDS and DNP in binary and 2.08 and 2.07 nm for (Hb–DNP) DIDS and (Hb–DIDS) DNP complexes in ternary systems, respectively. Time-resolved fluorescence spectroscopy confirmed static quenching for Hb in the presence of DIDS and DNP in both systems. Furthermore, an increase in ellipticity values of Hb upon interaction with DIDS and DNP showed secondary structural changes of protein that determine to disrupt of hydrogen bonds and electrostatic interactions. Our results showed that the Hb destabilize in the presence of DIDS and DNP. Molecular modeling of the possible binding sites of DIDS and DNP in binary and ternary systems in Hb confirmed the experimental results.     

Evaluation of DNA,BSA binding,and antimicrobial activity of new synthesized neodymium complex containing 29-dimethyl 110-phenanthroline

In order to evaluate biological potential of a novel synthesized complex [Nd(dmp)₂Cl₃.OH₂] where dmp is 29-dimethyl 110-phenanthroline, the DNA-binding, cleavage, BSA binding, and antimicrobial activity properties of the complex are investigated by multispectroscopic techniques study in physiological buffer (pH 7.2).The intrinsic binding constant (K_b) for interaction of Nd(III) complex and FS–DNA is calculated by UV–Vis (K_b = 2.7 ± 0.07 × 10⁵) and fluorescence spectroscopy (K_b = 1.13 ± 0.03 × 10⁵). The Stern–Volmer constant (K_SV), thermodynamic parameters including free energy change (ΔG°), enthalpy change (?H°), and entropy change (?S°), are calculated by fluorescent data and Vant’ Hoff equation. The experimental results show that the complex can bind to FS–DNA and the major binding mode is groove binding. Meanwhile, the interaction of Nd(III) complex with protein, bovine serum albumin (BSA), has also been studied by using absorption and emission spectroscopic tools. The experimental results show that the complex exhibits good binding propensity to BSA. The positive ΔH° and ?S° values indicate that the hydrophobic interaction is main force in the binding of the Nd(III) complex to BSA, and the complex can quench the intrinsic fluorescence of BSA remarkably through a static quenching process. Also, DNA cleavage was investigated by agarose gel electrophoresis that according to the results cleavage of DNA increased with increasing of concentration of the complex. Antimicrobial screening test gives good results in the presence of Nd(III) complex system.     

Biophysical and computational comparison on the binding affinity of three important nutrients to β-lactoglobulin: folic acid,ascorbic acid and vitamin K3

Small globular protein, β-lactoglobulin (βLG), which has significant affinity toward many drugs, is the most abundant whey protein in milk. In this study, the interaction of βLG with three important nutrients, ascorbic acid (ASC), folic acid (FOL), and vitamin K3 (VK3) was investigated by spectroscopic methods (UV–visible and fluorescence) along with molecular docking technique. The results of fluorescence measurements showed that studied nutrients strongly quenched βLG fluorescence in static (FOL and ACS) or static–dynamic combined quenching (VK3) mode. The values of binding constants (K_βLG-ASC ~ 4.34 × 10⁴ M^?1, K_βLG-FOL ~ 1.67 × 10⁴ M^?1and K_βLG-VK3 ~ 13.49 × 10⁴ M^?1 at 310 K) suggested that VK3 and FOL had stronger binding affinity toward βLG than ASC. Thermodynamic analysis indicated that hydrophobic interactions are the major forces in the stability of FOL–βLG complex with enthalpy- and entropy-driving mode while, hydrogen bonds and van der Waals interactions play a major role for βLG–ASC and βLG–VK3 associations. The results of 3D fluorescence FT-IR and UV–Visible measurements indicated that the binding of above nutrients to βLG may induce conformational and micro-environmental changes of protein. Also, there is a reciprocal complement between spectroscopic techniques and molecular docking modeling. The docking results indicate that the ASC, FOL, and VK3 bind to residues located in the subdomain B of βLG. Finally, this report suggests that βLG could be used as an effective carrier of above nutrients in functional foods.     

Investigation of the interaction between salvianolic acid C and xanthine oxidase: Insights from experimental studies merging with molecular docking methods

Xanthine oxidase (XO) has emerged as an important target for gout. In our previous study, salvianolic acid C (SAC) was found to show potent XO inhibitory activity, whereas the interaction mechanism was still not clear. Herein, an integrated approach consisting of enzyme kinetics, multi-spectroscopic methods and molecular docking was employed to investigate the interaction between SAC and XO. Consequently, SAC exhibited a rapid and mixed-type inhibition of XO with IC₅₀ of 5.84 ± 0.18 μM. The fluorescence data confirmed that SAC presented a strong fluorescence quenching effect through a static quenching procedure. The values of enthalpy change, entropy change and Gibbs free energy change indicated that their binding was spontaneous and driven mainly by hydrophobic interactions. Analysis of synchronous fluorescence, circular dichroism and fourier transform infrared spectra demonstrated that SAC induced conformational changes of the enzyme. Besides, further molecular docking revealed that SAC occupied the catalytic center resulting in the inhibition of XO activity. This study provides a comprehensive understanding on the interaction mechanism of SAC on XO.     

Conformational dynamics of DnaB helicase upon DNA and nucleotide binding: analysis by intrinsic tryptophan fluorescence quenching

DnaB helicase of E. coli unwinds duplex DNA in the replication fork using the energy of ATP hydrolysis. We have analyzed structural and conformational changes in the DnaB protein in various nucleotides and DNA bound intermediate states by fluorescence quenching analysis of intrinsic fluorescence of native tryptophan (Trp) residues in DnaB. Fluorescence quenching analysis indicated that Trp48 in domain alpha is in a hydrophobic environment and resistant to fluorescence quenchers such as potassium iodide (KI). In domain beta, Trp294 was found to be in a partially hydrophobic environment, whereas Trp456 in domain gamma appeared to be in the least hydrophobic environment. Binding of oligonucleotides to DnaB helicase resulted in a significant attenuation of the fluorescence quenching profile, indicating a change in conformation. ATPgammaS or ATP binding appeared to lead to a conformation in which Trp residues had a higher degree of solvent exposure and fluorescence quenching. However, the most dramatic increase of Trp fluorescence quenching was observed with ADP binding with a possible conformational relaxation. Site-specific Trp --> Cys mutants of DnaB helicase demonstrated that conformational change upon ADP binding could be attributed exclusively to a conformational transition in the alpha domain leading to an increase in the solvent exposure of Trp48. However, formation of DnaB.ATPgammaS.DNA ternary complex led to a conformation with a fluorescence quenching profile similar to that observed with DnaB alone. The DnaB.ADP.DNA ternary complex produced a quenching curve similar to that of DnaB.ADP complex pointing to a change in conformation due to ATP hydrolysis. There are at least four identifiable structural/conformational states of DnaB helicase that are likely important in the helicase activity. The noncatalytic alpha domain in the N-terminus appeared to undergo the most significant conformational changes during nucleotide binding and hydrolysis. This is the first reported elucidation of the putative role of domain alpha, which is essential for DNA helicase action. We have correlated these results with partial structural models of alpha, beta, and gamma domains     

Hydrogen bonding-assisted interaction between amitriptyline hydrochloride and hemoglobin: spectroscopic and molecular dynamics studies

Herein, we have explored the interaction between amitriptyline hydrochloride (AMT) and hemoglobin (Hb), using steady-state and time-resolved fluorescence spectroscopy, UV–visible spectroscopy, and circular dichroism spectroscopy, in combination with molecular docking and molecular dynamic (MD) simulation methods. The steady-state fluorescence reveals the static quenching mechanism in the interaction system, which was further confirmed by UV–visible and time-resolved fluorescence spectroscopy. The binding constant, number of binding sites, and thermodynamic parameters viz. ΔG, ΔH, ΔS are also considered; result confirms that the binding of the AMT with Hb is a spontaneous process, involving hydrogen bonding and van der Waals interactions with a single binding site, as also confirmed by molecular docking study. Synchronous fluorescence, CD data, and MD simulation results contribute toward understanding the effect of AMT on Hb to interpret the conformational change in Hb upon binding in aqueous solution.     

Insight into the interaction of antitubercular and anticancer compound clofazimine with human serum albumin: spectroscopy and molecular modelling

The binding of clofazimine to human serum albumin (HSA) was investigated by applying optical spectroscopy and molecular docking methods. Fluorescence quenching data revealed that clofazimine binds to protein with binding constant in the order of 10⁴ M^?1, and with the increase in temperature, Stern–Volmer quenching constants gradually decreased indicating quenching mode to be static. The UV–visible spectra showed increase in absorbance upon interaction of HSA with clofazimine which further reveals formation of the drug–albumin complex. Thermodynamic parameters obtained from fluorescence data indicate that the process is exothermic and spontaneous. Forster distance (R_o) obtained from fluorescence resonance energy transfer is found to be 2.05 nm. Clofazimine impelled rise in α-helical structure in HSA as observed from far-UV CD spectra while there are minor alterations in tertiary structure of the protein. Clofazimine interacts strongly with HSA inducing secondary structure in the protein and slight alterations in protein topology as suggested by dynamic light scattering results. Moreover, docking results indicate that clofazimine binds to hydrophobic pocket near to the drug site II in HSA.