Analysis of tryptophan fluorescence lifetimes in a series of human serum albumin mutants with substitutions in subdomain 2 A

Tryptophan 214, the only tryptophan residue in human serum albumin, is located in the physiologically important subdomain 2A ligand binding site. In the present study the fluorescence lifetime of tryptophan 214 in the following human serum albumin (HSA) mutants with substitutions in subdomain 2A were determined: K195M, K199M, F211V, R218M, R218H, R218A, R222M, H242V, and R257M. An HSA mutant in which tryptophan was moved from subdomain 2A to subdomain 3A (W214L/Y411W) was also examined. Additionally, the fluorescence lifetime of tryptophan 214 in an HSA fragment consisting of subdomains 1A, 1B, and 2A (1A-1B-2A HSA) was determined. For those species expected to have the most dramatic changes in tryptophan microenvironment, W214L/Y411W and 1A-1B-2A HSA, clear changes in tryptophan lifetimes were observed. Significant changes were also seen for those species with mutations at position 218, which is next to tryptophan in the X-ray structure of HSA. However, significant changes were also observed for H242V and R257M, which contain substitutions at positions not immediately adjacent to tryptophan 214, highlighting the conformational flexibility of subdomain 2A.     

A method of chemiluminescence coupled with ultrafiltration for investigating the interaction between ibuprofen and human serum albumin

In acidic media, ibuprofen substantially enhanced the weak chemiluminescence (CL) produced by sodium sulfite and potassium permanganate. The increased signals were linearly correlated with ibuprofen concentrations ranging from 1.2 × 10^‐3 to 4.8 μM, with a detection limit of 4.8 × 10^‐4 μM. Two ultrafiltration (UF) membranes were used to construct a unit for trapping 0.15 and 0.75 μM human serum albumin (HSA) and coupled online with the CL system. At low HSA concentrations, the numbers of bound molecules per binding site were calculated to be 0.9 for Sudlow site I and 6.2 for Sudlow site II. The association constants on these binding sites were 5.9 × 10⁵ and 3.4 × 10⁴ M^‐1, respectively. Our CL–UF protocol presents a rapid and sensitive method for studies on drug–protein interaction. Copyright © 2012 John Wiley & Sons, Ltd.     

Interaction of a tyrosine kinase inhibitor,vandetanib with human serum albumin as studied by fluorescence quenching and molecular docking

Interaction of a tyrosine kinase inhibitor, vandetanib (VDB), with the major transport protein in the human blood circulation, human serum albumin (HSA), was investigated using fluorescence spectroscopy, circular dichroism (CD) spectroscopy, and molecular docking analysis. The binding constant of the VDB–HSA system, as determined by fluorescence quenching titration method was found in the range, 8.92–6.89?×?10^3?M^?1 at three different temperatures, suggesting moderate binding affinity. Furthermore, decrease in the binding constant with increasing temperature revealed involvement of static quenching mechanism, thus affirming the formation of the VDB–HSA complex. Thermodynamic analysis of the binding reaction between VDB and HSA yielded positive ΔS (52.76 J?mol^?1 K^?1) and negative ΔH (?6.57?kJ?mol^?1) values, which suggested involvement of hydrophobic interactions and hydrogen bonding in stabilizing the VDB–HSA complex. Far-UV and near-UV CD spectral results suggested alterations in both secondary and tertiary structures of HSA upon VDB-binding. Three-dimensional fluorescence spectral results also showed significant microenvironmental changes around the Trp residue of HSA consequent to the complex formation. Use of site-specific marker ligands, such as phenylbutazone (site I marker) and diazepam (site II marker) in competitive ligand displacement experiments indicated location of the VDB binding site on HSA as Sudlow’s site I (subdomain IIA), which was further established by molecular docking results. Presence of some common metal ions, such as Ca²⁺, Zn²⁺, Cu²⁺, Ba²⁺, Mg²⁺, and Mn²⁺ in the reaction mixture produced smaller but significant alterations in the binding affinity of VDB to HSA.     

Deciphering the binding of carbendazim (fungicide) with human serum albumin: A multi-spectroscopic and molecular modelling studies

Carbendazim is a benzimidazole fungicide used to control the fungal invasion. However, its exposure might lead to potential health problems. The present study evaluates the interaction of carbendazim (CAR) with human serum albumin (HSA) which is an important drug carrier protein and plays a very crucial role in the transportation of small molecules. A number of biophysical techniques were employed to investigate the binding of CAR with HSA. The increased UV-absorption of HSA on titrating with CAR suggests the formation of HSA–CAR complex and it could be due to the exposure of aromatic residues. The fluorescence study confirmed that CAR quenches the fluorescence of HSA and showed the static mode of quenching. CAR (50 µM) quenches around 56.14% of the HSA fluorescence. The quenching constant, binding constant, number of binding site and free energy change was calculated by fluorescence quenching experiment. Competitive displacement assay showed Sudlow’s site I as the primary binding site of CAR on HSA. The synchronous fluorescence study revealed the perturbation in the microenvironment around tyrosine and tryptophan residues upon binding of CAR to HSA. The circular dichroism results suggested that the binding of CAR to HSA altered its secondary structure. Molecular docking experiment demonstrated the binding of CAR to Sudlow’s site I of HSA. Docking studies suggested that the hydrogen bonding, van der Waals and pi-alkyl are playing role in the interaction of CAR with HSA. The study confirmed the conformational changes within HSA upon binding of CAR.     

Probing the structure of the warfarin-binding site on human serum albumin using site-directed mutagenesis

The binding of warfarin to the following human serum albumin (HSA) mutants was examined: K195M, K199M, F211V, W214L, R218M, R222M, H242V, and R257M. Warfarin bound to human serum albumin (HSA) exhibits an intrinsic fluorescence that is approximately 10-fold greater than the corresponding signal for warfarin in aqueous solution. This property of the warfarin/HSA complex has been widely used to determine the dissociation constant for the interaction. In the present study, such a technique was used to show that specific substitutions in subdomain 2A altered the affinity of HSA for warfarin. The fluorescence of warfarin/mutant HSA complexes varied widely from the fluorescence of the warfarin/wild-type HSA complex at pH = 7.4, suggesting changes in the structure of the complex resulting from specific substitutions. The fluorescence of the warfarin/wild-type HSA complex increases about twofold as the pH is increased from 6.0 to 9.0 due to the neutral-to-base (N-B) transition, a conformational change that occurs in HSA as a function of pH. Changes in the fluorescence of warfarin/mutant HSA complexes as a function of pH suggests novel behavior for most HSA species examined. For the HSA mutants F211V and H242V, the midpoint of the N-B transition shifts from a wild-type pH of 7.8 to a pH value of 7.1-7.2.     

A fluorescent fatty acid probe, DAUDA, selectively displaces two myristates bound in human serum albumin

11-(Dansylamino) undecanoic acid (DAUDA) is a dansyl-type fluorophore and has widely used as a probe to determine the binding site for human serum albumin (HSA). Here, we reported that structure of HSA-Myristate-DAUDA ternary complex and identified clearly the presence of two DAUDA molecules at fatty acid (FA) binding site 6 and 7 of HSA, thus showing these two sites are weak FA binding sites. This result also show that DAUDA is an appropriate probe for FA site 6 and 7 on HSA as previous studied, but not a good probe of FA binding site 1 that is likely bilirubin binding site on HSA.     

Probing the interaction of thionine with human serum albumin by multispectroscopic studies and its in vitro cytotoxic activity toward MCF-7 breast cancer cells

The studies on protein–dye interactions are important in biological process and it is regarded as vital step in rational drug design. The interaction of thionine (TH) with human serum albumin (HSA) was analyzed using isothermal titration calorimetry (ITC), spectroscopic, and molecular docking technique. The emission spectral titration of HSA with TH revealed the formation of HSA–TH complex via static quenching process. The results obtained from absorption, synchronous emission, circular dichroism, and three-dimensional (3D) emission spectral studies demonstrated that TH induces changes in the microenvironment and secondary structure of HSA. Results from ITC experiments suggested that the binding of TH dye was favored by negative enthalpy and a favorable entropy contribution. Site marker competitive binding experiments revealed that the binding site of TH was located in subdomain IIA (Sudlow site I) of HSA. Molecular docking study further substantiates that TH binds to the hydrophobic cavity of subdomain IIA (Sudlow site I) of HSA. Further, we have studied the cytotoxic activity of TH and TH–HSA complex on breast cancer cell lines (MCF-7) by MTT assay and LDH assay. These studies revealed that TH–HSA complex showed the higher level of cytotoxicity in cancer cells than TH dye-treated MCF-7 cells and the significant adverse effect did not found in the normal HBL-100 cells. Fluorescence microscopy analyses of nuclear fragmentation studies validate the significant reduction of viability of TH–HSA-treated human MCF-7 breast cancer cells through activation of apoptotic-mediated pathways.     

Is the Sudlow site I of human serum albumin more generous to adopt prospective anti-cancer bioorganic compound than that of bovine: A combined spectroscopic and docking simulation approach

A comparative biophysical study on the individual conformational adaptation embraced by two homologous serum albumins (SA) (bovine and human) towards a potential anticancer bioorganic compound 2-(6-chlorobenzo[d] thiazol-2-yl)-1H-benzo[de] isoquinoline-1,3(2H)- dione (CBIQD) is apparent from the discrimination in binding behavior and the ensuing consequences accomplished by combined in vitro optical spectroscopy, in silico molecular docking and molecular dynamics (MD) simulation. The Sudlow site I of HSA although anion receptive, harbors neutral CBIQD in Sudlow site I (subdomain IIA, close to Trp) of HSA, while in BSA its prefers to snugly fit into Sudlow site II (subdomain IIIA, close to Tyr). Based on discernable diminution of HSA mean fluorescence lifetime as a function of biluminophore concentration, facile occurrence of fluorescence resonance energy transfer (FRET) is substantiated as the probable quenching mechanism accompanied by structural deformations in the protein ensemble. CBIQD establishes itself within HSA close to Trp214, and consequently reduces the micropolarity of the cybotactic environment that is predominantly constituted by hydrophobic amino acid residues. The stronger association of CBIQD with HSA encourages an allosteric modulation leading to slight deformation in its secondary structure whereas for BSA the association is comparatively weaker. Sudlow site I of HSA is capable to embrace a favorable conformation like malleable gold to provide room for incoming CBIQD, whereas for BSA it behaves more like rigid cast-iron which does not admit any change thus forcing CBIQD to occupy an altogether different binding location i.e. the Sudlow site II. The anticancer CBIQD is found to be stable within the HSA scaffold as vindicated by root mean square deviation (RMSD) and root mean square fluctuation (RMSF) obtained by MD simulation. A competitively inhibited esterase-like activity of HSA upon CBIQD binding to Lys199 and Arg257 residues, plausibly envisions that similar naphthalimide based prodrugs, bearing ester functionality, can be particularly activated by Sudlow site I of HSA. The consolidated spectroscopic research described herein may encourage design of naphthalimide based pro-drugs for effective in vivo biodistribution using HSA-based drug delivery systems.     

Interactive association between RhoA transcriptional signaling inhibitor,CCG1423 and human serum albumin: Biophysical and in silico studies

Multiple spectroscopic techniques, such as fluorescence, absorption, and circular dichroism along with in silico studies were used to characterize the binding of a potent inhibitor molecule, CCG1423 to the major transport protein, human serum albumin (HSA). Fluorescence and absorption spectroscopic results confirmed CCG1423–HSA complex formation. A strong binding affinity stabilized the CCG1423–HSA complex, as evident from the values of the binding constant (K_a = 1.35 × 10⁶–5.43 × 10⁵ M^?1). The K_SV values for CCG1423–HSA system were inversely correlated with temperature, suggesting the involvement of static quenching mechanism. Thermodynamic data anticipated that CCG1423–HSA complexation was mainly driven by hydrophobic and van der Waals forces as well as hydrogen bonds. In silico analysis also supported these results. Three-dimensional fluorescence and circular dichroism spectral analysis suggested microenvironmental perturbations around protein fluorophores and structural (secondary and tertiary) changes in the protein upon CCG1423 binding. CCG1423 binding to HSA also showed some protection against thermal denaturation. Site-specific marker-induced displacement results revealed CCG1423 binding to Sudlow’s site I of HSA, which was also confirmed by the computational results. A few common ions were also found to interfere with the CCG1423–HSA interaction.     

Structural insights into human serum albumin-mediated prostaglandin catalysis

Previous studies have shown that many arachidonic acid metabolites bind to human serum albumin (HSA) and that the metabolism of these molecules is altered as a result of binding. The present study attempted to gain insights into the mechanisms by which prostaglandins bound to subdomain 2A of HSA are metabolized by catalytic processes. The breakdown of the prostaglandin 15-keto-PGE(2) to 15-keto-PGA(2) and 15-keto-PGB(2) in the presence of wild-type HSA and a number of subdomain 2A mutants was examined using a previously validated spectroscopic method which monitors absorbance at 505 nm. The species examined using this method were wild-type HSA, K195M, K199M, F211V, W214L, R218M, R218P, R218H, R222M, H242V, R257M, and bovine serum albumin. Previous studies of HSA-mediated catalysis indicated that the breakdown of HSA-bound prostaglandins results from an alkaline microenvironment in the binding site. Our results show that the catalytic breakdown of HSA-bound 15-keto-PGE(2) to 15-keto-PGB(2) results from two specific processes which are modulated by specific amino acid residues. Specifically, some amino acid residues modulate the rate of step 1, the conversion of 15-keto-PGE(2) to 15-keto-PGA(2), while other residues modulate the rate of step 2, the conversion of 15-keto-PGA(2) to 15-keto-PGB(2). Some residues modulate the rate of steps 1 and 2. In total, while our results support the involvement of certain basic amino acid residues in the catabolism of HSA-bound 15-keto-PGE(2), our data suggest that metabolism of HSA-bound prostaglandins may be a more complex and specific process than previously thought.     

Probing the interaction of anticancer drug temsirolimus with human serum albumin: molecular docking and spectroscopic insight

The binding interaction between temsirolimus, an important antirenal cancer drug, and HSA, an important carrier protein was scrutinized making use of UV and fluorescence spectroscopy. Hyper chromaticity observed in UV spectroscopy in the presence of temsirolimus as compared to free HSA suggests the formation of complex between HSA and temsirolimus. Fluorescence quenching experiments clearly showed quenching in the fluorescence of HSA in the presence of temsirolimus confirming the complex formation and also confirmed that static mode of interaction is operative for this binding process. Binding constant values obtained through UV and fluorescence spectroscopy reveal strong interaction; temsirolimus binds to HSA at 298 K with a binding constant of 2.9 × 10⁴ M^?1implying the strength of interaction. The negative Gibbs free energy obtained through Isothermal titration calorimetry as well as quenching experiments suggests that binding process is spontaneous. Molecular docking further provides an insight of various residues that are involved in this binding process; showing the binding energy to be -12.9 kcal/mol. CD spectroscopy was retorted to analyze changes in secondary structure of HSA; increased intensity in presence of temsirolimus showing changes in secondary structure of HSA induced by temsirolimus. This study is of importance as it provides an insight into the binding mechanism of an important antirenal cancer drug with an important carrier protein. Once temsirolimus binds to HSA, it changes conformation of HSA which in turn can alter the functionality of this important carrier protein and this altered functionality of HSA can be highlighted in variety of diseases.     

Fipronil recognition by the FA1 site of human serum albumin

Fipronil is a broad‐spectrum pesticide widely used in agriculture, horticulture, and forestry. Because fipronil can cause a variety of toxic effects in animals and humans, its use is authorized as a pesticide in veterinary medicinal products for pets, but not for the treatment of livestock animals whose products are intended for consumption. Recently, however, the presence of fipronil residues has been detected in the eggs and meat of layer hens from farms located in different European countries. Given the relevance of fipronil toxicity for human health, it is important to gain information concerning its fate in the human body, including its binding mode to human serum albumin (HSA), the most abundant protein in plasma. Here, the inhibition of heme‐Fe(III) binding to the fatty acid site 1 (FA1) of HSA by fipronil is reported. Docking simulations support functional data, indicating that the FA1 site is the preferential cleft for fipronil recognition by HSA. The affinity of fipronil for HSA (K_f = 1.9 × 10^?6 M, at pH 7.3, and 20.0°C) may be relevant in vivo. Indeed, HSA could play a pivotal role in fipronil transport and scavenging, thus reducing the pesticide‐free plasmatic levels, with consequent reduced systemic toxicity. In turn, fipronil binding to the FA1 site of HSA could impair the recognition of endogenous and exogenous molecules.     

Extending the half-life of a fab fragment through generation of a humanized anti-human serum albumin Fv domain: An investigation into the correlation between affinity and serum half-life

We generated an anti-albumin antibody, CA645, to link its Fv domain to an antigen-binding fragment (Fab), thereby extending the serum half-life of the Fab. CA645 was demonstrated to bind human, cynomolgus, and mouse serum albumin with similar affinity (1–7 nM), and to bind human serum albumin (HSA) when it is in complex with common known ligands. Importantly for half-life extension, CA645 binds HSA with similar affinity within the physiologically relevant range of pH 5.0 – pH 7.4, and does not have a deleterious effect on the binding of HSA to neonatal Fc receptor (FcRn). A crystal structure of humanized CA645 Fab in complex with HSA was solved and showed that CA645 Fab binds to domain II of HSA. Superimposition with the crystal structure of FcRn bound to HSA confirmed that CA645 does not block HSA binding to FcRn. In mice, the serum half-life of humanized CA645 Fab is 84.2 h. This is a significant extension in comparison with < 1 h for a non-HSA binding CA645 Fab variant. The Fab-HSA structure was used to design a series of mutants with reduced affinity to investigate the correlation between the affinity for albumin and serum half-life. Reduction in the affinity for MSA by 144-fold from 2.2 nM to 316 nM had no effect on serum half-life. Strikingly, despite a reduction in affinity to 62 µM, an extension in serum half-life of 26.4 h was still obtained. CA645 Fab and the CA645 Fab-HSA complex have been deposited in the Protein Data Bank (PDB) with accession codes, 5FUZ and 5FUO, respectively.     

Multi-spectroscopic and molecular modelling approach to investigate the interaction of riboflavin with human serum albumin

Riboflavin (RF) plays an important role in various metabolic redox reactions in the form of flavin adenine dinucleotide and flavin mononucleotide. Human serum albumin (HSA) is an important protein involved in the transportation of drugs, hormones, fatty acid and other molecules which determine the biodistribution and physiological fate of these molecules. In this study, we have investigated the interaction of riboflavin RF with HSA under simulative physiological conditions using various biophysical, calorimetric and molecular docking techniques. Results demonstrate the formation of riboflavin–HSA complex with binding constant in the order of 10⁴ M^?1. Fluorescence spectroscopy confirms intermediate strength having a static mode of quenching with stoichiometry of 1:1. Experimental results suggest that the binding site of riboflavin mainly resides in sub-domain IIA of HSA and that ligand interaction increases the α-helical content of HSA. These parameters were further verified by isothermal titration calorimetry ITC which confirms the thermodynamic parameters obtained by fluorescence spectroscopy. Molecular docking was employed to suggest a binding model. Based on thermodynamic, spectroscopic and computational observations it can be concluded that HSA-riboflavin complex is mainly stabilized by various non-covalent forces with binding energy of ?7.2 kcal mol^?1.     

Chromatographic analysis of acetohexamide binding to glycated human serum albumin

Acetohexamide is a drug used to treat type II diabetes and is tightly bound to the protein human serum albumin (HSA) in the circulation. It has been proposed that the binding of some drugs with HSA can be affected by the non-enzymatic glycation of this protein. This study used high-performance affinity chromatography to examine the changes in acetohexamide–HSA binding that take place as the glycation of HSA is increased. It was found in frontal analysis experiments that the binding of acetohexamide to glycated HSA could be described by a two-site model involving both strong and weak affinity interactions. The average association equilibrium constant (K_a) for the high affinity interactions was in the range of 1.2–2.0 × 10⁵ M⁻¹ and increased in moving from normal HSA to HSA with glycation levels that might be found in advanced diabetes. It was found through competition studies that acetohexamide was binding at both Sudlow sites I and II on the glycated HSA. The K_a for acetohexamide at Sudlow site I increased by 40% in going from normal HSA to minimally glycated HSA but then decreased back to near-normal values in going to more highly glycated HSA. At Sudlow site II, the K_a for acetohexamide first decreased by about 40% and then increased in going from normal HSA to minimally glycated HSA and more highly glycated HSA. This information demonstrates the importance of conducting both frontal analysis and site-specific binding studies in examining the effects of glycation on the interactions of a drug with HSA.     

Glycation of human serum albumin alters its binding efficacy towards the dietary polyphenols: a comparative approach

Diabetes is a major problem in the world. The proteins became modified during glycation after reacting with the reducing sugars (e.g. D-glucose) via non-enzymatic pathways. The glycated analogue of human serum albumin (HSA) has been characterized with the help of multi-spectroscopic methods. It has been observed that six glucose molecules can bind covalently to HSA under experimental condition. The binding affinity of the modified HSA towards the dietary polyphenols has been estimated using UV–vis and fluorescence spectroscopic techniques. The binding constant values of the ligands were found to decrease after the modification of HSA.     

Study of the Enantioselective Interaction of Diclofop and Human Serum Albumin by Spectroscopic and Molecular Modeling Approaches In Vitro

In this contribution, the enantioselective interactions between diclofop (DC) and human serum albumin (HSA) were explored by steady‐state and 3D fluorescence, ultraviolet‐visible spectroscopy (UV‐vis), and molecular modeling. The binding constants between R‐DC and HSA were 0.9213 × 10⁵, 0.9118 × 10⁵, and 0.9009 × 10⁵ L · mol^‐1 at 293, 303, 313 K, respectively. Moreover, the binding constants of S‐DC for HSA were 1.4766 × 10⁵, 1.2899 × 10⁵, and 1.0882 × 10⁵ L · mol^‐1 at 293, 303, and 313 K individually. Such consequences markedly implied the binding between DC enantiomers and HSA were enantioselective with higher affinity for S‐DC. Steady‐state fluorescence study evidenced the formation of DC‐HSA complex and there was a single class of binding site on HSA. The thermodynamic parameters (ΔH, ΔS, ΔG) of the reaction clearly indicated that hydrophobic effects and H‐bonds contribute to the formation of DC‐HSA complex, which was in excellent agreement with molecular simulations. In addition, both site‐competitive replacement and molecular modeling suggested that DC enantiomers were located within the binding pocket of Sudlow's site II. Furthermore, the alterations of HSA secondary structure in the presence of DC enantiomers were verified by UV‐vis absorption and 3D fluorescence spectroscopy. This study can provide important insight into the enantioselective interaction of physiological protein HSA with chiral aryloxyphenoxy propionate herbicides and gives support to the use of HSA for chiral pesticides ecotoxicology and environmental risk assessment. Chirality 25:719–725, 2013. © 2013 Wiley Periodicals, Inc.     

Cytotoxicity and comparative binding mechanism of piperine with human serum albumin and α-1-acid glycoprotein

Human serum albumin (HSA) and α-1-acid glycoprotein (AGP) (acute phase protein) are the plasma proteins in blood system which transports many drugs. To understand the pharmacological importance of piperine molecule, here, we studied the anti-inflammatory activity of piperine on mouse macrophages (RAW 264.7) cell lines, which reveals that piperine caused an increase in inhibition growth of inflammated macrophages. Further, the fluorescence maximum quenching of proteins were observed upon binding of piperine to HSA and AGP through a static quenching mechanism. The binding constants obtained from fluorescence emission were found to be K_piperine?=?5.7 ± .2 × 10⁵ M^?1 and K_piperine = 9.3± .25 × 10⁴ M^?1 which correspond to the free energy of ?7.8 and ?6.71 kcal M^?1at 25 °C for HSA and AGP, respectively. Further, circular dichrosim studies revealed that there is a marginal change in the secondary structural content of HSA due to partial destabilization of HSA–piperine complexes. Consequently, inference drawn from the site-specific markers (phenylbutazone, site I marker) studies to identify the binding site of HSA noticed that piperine binds at site I (IIA), which was further authenticated by molecular docking and molecular dynamic (MD) studies. The binding constants and free energy corresponding to experimental and computational analysis suggest that there are hydrophobic and hydrophilic interactions when piperine binds to HSA. Additionally, the MD studies have showed that HSA–piperine complex reaches equilibration state at around 3 ns, which prove that the HSA–piperine complex is stable in nature.     

Molecular Modeling of Cloned <Emphasis Type="Italic">Bacillus subtilis</Emphasis> Keratinase and Its Insinuation in Psoriasis Treatment Using Docking Studies

Present study demonstrated the expression of cloned Bacillus subtilis RSE163 keratinase gene and in silico binding affinities of deduced protein with psoriasis topical drugs for systemic absorption and permeation through skin. The ker gene expressed in E. coli showed significantly higher keratinase activity 450 ± 10.43 U representing 1342 bp nucleotides encoding 447 amino acids with molecular weight of 46 kDa. The modeled structure was validated using ramachandran’s plot showing 305 residues (84.3%) in most favoured region. Docking studies using extra precision (XP) method of Glide showed optimum binding affinities with the drugs Acitretin (? 39.62 kcal/mol), Clobetasol propionate (? 37.90 kcal/mol), Fluticasone (? 38.53 kcal/mol), Desonide (? 32.23 kcal/mol), Anthralin (? 38.04 kcal/mol), Calcipotreine (? 21.55 kcal/mol) and Mometasone (? 28.40 kcal/mol) in comparison to other psoriasis drugs. The results can further be correlated with in vitro enzymatic experiments using keratinase as an effective drug mediator through skin to serve the unmet need of industries.     

Experimental and computational studies on the binding of diazinon to human serum albumin

In the present research, the binding properties of diazinon (DZN), as an organophosphorus herbicide, to human serum albumin (HSA) were investigated using combination of spectroscopic, electrochemistry, and molecular modeling techniques. Changes in the UV–Vis and FT-IR spectra were observed upon ligand binding along with a significant degree of tryptophan fluorescence quenching on complex formation. The obtained results from spectroscopic and electrochemistry experiments along with the computational studies suggest that DZN binds to residues located in subdomains IIA of HSA with binding constant about 1410.9 M^?1 at 300 K. From the thermodynamic parameters calculated according to the van’t Hoff equation, the enthalpy change ΔH° and entropy change ΔS° were found to be ?16.695 and 0.116 KJ/mol K, respectively. The primary binding pattern is determined by hydrophobic interaction and hydrogen binding occurring in so-called site I of HSA. DZN could slightly alter the secondary structure of HSA. All of experimental results are supported by computational techniques such as docking and molecular dynamics simulation using a HSA crystal model.