Integrated spectroscopic and computational analyses unravel the molecular interaction of pesticide azinphos-methyl with bovine beta-lactoglobulin.

Organophosphorus are typically hazardous chemicals used in the pharmaceutical, agricultural, and other industries. They pose a serious risk to human life and can be fatal upon direct exposure. Hence, studying the interaction between such compounds with proteins is crucial for environmental, health, and food safety. In this study, we investigated the interaction mechanism between azinphos-methyl (AZM) and ß-lactoglobulin (BLG) at pH 7.4 using a combination of biophysical techniques. Intrinsic fluorescence investigations revealed that BLG fluorescence was quenched in the presence of increasing AZM concentrations. The quenching mechanism was identified as static, as evidenced by a decrease in the fluorescence quenching constant (1.25 × 104, 1.18 × 104, and 0.86 × 104 M-1) with an increase in temperatures. Thermodynamic calculations (ΔH > 0; ΔS > 0) affirmed the formation of a complex between AZM and BLG through hydrophobic interactions. The BLG's secondary structure was found to be increased due to AZM interaction. Ultraviolet -visible spectroscopy data showed alterations in BLG conformation in the presence of AZM. Molecular docking highlighted the significant role of hydrophobic interactions involving residues such as Val43, Ile56, Ile71, Val92, Phe105, and Met107 in the binding between BLG and AZM. A docking energy of -6.9 kcal mol-1, and binding affinity of 1.15 × 105 M-1 suggest spontaneous interaction between AZM and BLG with moderate to high affinity. These findings underscore the potential health risks associated with the entry of AZM into the food chain, emphasizing the need for further consideration of its impact on human health.

Agitation does not induce fibrillation in reduced hen egg-white lysozyme at physiological temperature and pH.

Several proteins and peptides tend to form an amyloid fibril, causing a range of unrelated diseases, from neurodegenerative to certain types of cancer. In the native state, these proteins are folded and soluble. However, these proteins acquired ß-sheet amyloid fibril due to unfolding and aggregation. The conversion mechanism from well-folded soluble into amorphous or amyloid fibril is not well understood yet. Here, we induced unfolding and aggregation of hen egg-white lysozyme (HEWL) by reducing agent dithiothreitol and applied mechanical sheering force by constant shaking (1000 rpm) on the thermostat for 7 days. Our turbidity results showed that reduced HEWL rapidly formed aggregates, and a plateau was attained in nearly 5 h of incubation in both shaking and non-shaking conditions. The turbidity was lower in the shaking condition than in the non-shaking condition. The thioflavin T binding and transmission electron micrographs showed that reduced HEWL formed amorphous aggregates in both conditions. Far-UV circular dichroism results showed that reduced HEWL lost nearly all alpha-helical structure, and ß-sheet secondary structure was not formed in both conditions. All the spectroscopic and microscopic results showed that reduced HEWL formed amorphous aggregates under both conditions.

Screening and identification of potential MERS-CoV papain-like protease (PLpro) inhibitors; Steady-state kinetic and Molecular dynamic studies.

MERS-CoV belongs to the coronavirus group. Recent years have seen a rash of coronavirus epidemics. In June 2012, MERS-CoV was discovered in the Kingdom of Saudi Arabia, with 2,591 MERSA cases confirmed by lab tests by the end of August 2022 and 894 deaths at a case-fatality ratio (CFR) of 34.5% documented worldwide. Saudi Arabia reported the majority of these cases, with 2,184 cases and 813 deaths (CFR: 37.2%), necessitating a thorough understanding of the molecular machinery of MERS-CoV. To develop antiviral medicines, illustrative investigation of the protein in coronavirus subunits are required to increase our understanding of the subject. In this study, recombinant expression and purification of MERS-CoV (PLpro), a primary goal for the development of 22 new inhibitors, were completed using a high throughput screening methodology that employed fragment-based libraries in conjunction with structure-based virtual screening. Compounds 2, 7, and 20, showed significant biological activity. Moreover, a docking analysis revealed that the three compounds had favorable binding mood and binding free energy. Molecular dynamic simulation demonstrated the stability of compound 2 (2-((Benzimidazol-2-yl) thio)-1-arylethan-1-ones) the strongest inhibitory activity against the PLpro enzyme. In addition, disubstitutions at the meta and para locations are the only substitutions that may boost the inhibitory action against PLpro. Compound 2 was chosen as a MERS-CoV PLpro inhibitor after passing absorption, distribution, metabolism, and excretion studies; however, further investigations are required.

Liposome-Mediated Delivery of MERS Antigen Induces Potent Humoral and Cell-Mediated Immune Response in Mice.

The advancements in the field of nanotechnology have provided a great platform for the development of effective antiviral vaccines. Liposome-mediated delivery of antigens has been shown to induce the antigen-specific stimulation of the humoral and cell-mediated immune responses. Here, we prepared dried, reconstituted vesicles (DRVs) from DPPC liposomes and used them as the vaccine carrier system for the Middle East respiratory syndrome coronavirus papain-like protease (DRVs-MERS-CoV PLpro). MERS-CoV PLpro emulsified in the Incomplete Freund's Adjuvant (IFA-MERS-CoV PLpro) was used as a control. Immunization of mice with DRVs-MERS-CoV PLpro did not induce any notable toxicity, as revealed by the levels of the serum alanine transaminase (ALT), aspartate transaminase (AST), blood urea nitrogen (BUN) and lactate dehydrogenase (LDH) in the blood of immunized mice. Immunization with DRVs-MERS-CoV PLpro induced greater antigen-specific antibody titer and switching of IgG1 isotyping to IgG2a as compared to immunization with IFA-MERS-CoV PLpro. Moreover, splenocytes from mice immunized with DRVs-MERS-CoV PLpro exhibited greater proliferation in response to antigen stimulation. Moreover, splenocytes from DRVs-MERS-CoV PLpro-immunized mice secreted significantly higher IFN-γ as compared to splenocytes from IFA-MERS-CoV PLpro mice. In summary, DRVs-MERS-CoV PLpro may prove to be an effective prophylactic formulation to prevent MERS-CoV infection.

Quinoline yellow dye stimulates whey protein fibrillation via electrostatic and hydrophobic interaction: A biophysical study.

Amyloid fibril formation of proteins is associated with a number of neurodegenerative diseases. Several small molecules can accelerate the amyloid fibril formation in vitro and in vivo. However, the molecular mechanism of amyloid fibrillation is still unclear. In this study, we investigated how the food dye quinoline yellow (QY) induces amyloid fibrillation in α-lactalbumin (α-LA), a major whey protein, at pH 2.0. We used several spectroscopy techniques and a microscopy technique to explore how QY provokes amyloid fibrillation in α-LA. From turbidity and Rayleigh light scattering experiments, we found that QY promotes α-LA aggregation in a concentration-dependent manner; the optimal concentration for α-LA aggregation was 0.15 to 10.00 mM. Below 0.1 mM, no aggregation occurred. Quinoline yellow-induced aggregation was a rapid process that escaped the lag phase, but it depended on the concentrations of both α-LA and QY. We also demonstrated that aggregation switched the secondary structure of α-LA from α-helices to cross-ß-sheets. We then confirmed the amyloid-like structure of aggregated α-LA by transmission electron microscopy measurements. Molecular docking and simulation confirmed the stability of the α-LA-QY complex due to the formation of 1 hydrogen bond with Lys99 and 2 electrostatic interactions with Arg70 and Lys99, along with hydrophobic interactions with Leu59 and Tyr103. This study will aid in our understanding of how small molecules induce aggregation of proteins inside the stomach (low pH) and affect the digestive process.

Structural Refolding and Thermal Stability of Myoglobin in the Presence of Mixture of Crowders: Importance of Various Interactions for Protein Stabilization in Crowded Conditions.

The intracellular environment is overcrowded with a range of molecules (small and large), all of which influence protein conformation. As a result, understanding how proteins fold and stay functional in such crowded conditions is essential. Several in vitro experiments have looked into the effects of macromolecular crowding on different proteins. However, there are hardly any reports regarding small molecular crowders used alone and in mixtures to observe their effects on the structure and stability of the proteins, which mimics of the cellular conditions. Here we investigate the effect of different mixtures of crowders, ethylene glycol (EG) and its polymer polyethylene glycol (PEG 400 Da) on the structural and thermal stability of myoglobin (Mb). Our results show that monomer (EG) has no significant effect on the structure of Mb, while the polymer disrupts its structure and decreases its stability. Conversely, the additive effect of crowders showed structural refolding of the protein to some extent. Moreover, the calorimetric binding studies of the protein showed very weak interactions with the mixture of crowders. Usually, we can assume that soft interactions induce structural perturbations while exclusion volume effects stabilize the protein structure; therefore, we hypothesize that under in vivo crowded conditions, both phenomena occur and maintain the stability and function of proteins.

Molecular interaction of tea catechin with bovine ß-lactoglobulin: A spectroscopic and in silico studies.

Polyphenols has attained pronounced attention due to their beneficial values of health and found to prevent several chronic diseases. Here, we elucidated binding mechanism between frequently consumed polyphenol "tea catechin" and milk protein bovine beta-lactoglobulin (ß-Lg). We investigated the conformational changes of ß-Lg due to interaction with catechin using spectroscopic and in silico studies. Fluorescence quenching data (Stern-Volmer quenching constant) revealed that ß-Lg interacted with catechin via dynamic quenching. Thermodynamic data revealed that the interaction between ß-Lg and catechin is endothermic and spontaneously interacted mainly through hydrophobic interactions. The UV-Vis absorption and far-UV circular dichroism (CD) spectroscopy exhibited that the tertiary as well as secondary structure of ß-Lg distorted after interaction with catechin. Molecular docking and simulation studies also confirm that catechin binds at the central cavity of ß-Lg with high affinity (~105 M-1) and hydrophobic interactions play significant role in the formation of a stable ß-Lg-catechin complex.

Overexpression, purification and enzymatic characterization of a recombinant Arabian camel Camelus dromedarius glucose-6-phosphate dehydrogenase.

In a previous study the full-length open reading frame of the Arabian camel, Camelus dromedarius liver cytosolic glucose-6-phosphate dehydrogenase (G6PD) cDNA was determined using reverse transcription polymerase chain reaction. The C. dromedarius cDNA was found to be 1545 nucleotides (accession number JN098421) that encodes a protein of 515 amino acids residues. In the present study, C. dromedarius recombinant G6PD was heterologously overexpressed in Escherichia coli BL21 (DE3) pLysS and purified by immobilized metal affinity fast protein liquid chromatography (FPLC) in a single step. The purity and molecular weight of the enzyme were analyzed on SDS-PAGE and the purified enzyme showed a single band on the gel with a molecular weight of 63.0 KDa. The specific activity was determined to be 2000 EU/mg protein. The optimum temperature and pH were found to be 60 °C and 7.4, respectively. The isoelectric point (pI) for the purified G6PD was determined to be 6.4. The apparent Km values for the two substrates NADP+ and G6P were found to be 23.2 µM and 66.7 µM, respectively. The far-UV circular dichroism (CD) spectra of G6PD showed that it has two minima at 208 and 222 nm as well as maxima at 193 nm which is characteristic of high content of α-helix. Moreover, the far-UV CD spectra of the G6PD in the presence or absence of NADP+ were nearly identical.

Spectroscopic and thermodynamic properties of recombinant heat shock protein A6 from Camelus dromedarius.

Heat shock protein A6, also known as HSP70B', is a member of the Hsp70 family of molecular chaperones. Under stressed conditions, the level of HSPA6 increases substantially, and the protein has been targeted as a biomarker of cellular stress in several studies. We report the spectroscopic and thermodynamic properties of Arabian camel species cHSPA6, determined by measurement of intrinsic and extrinsic fluorescence emission, and use of far-UV circular dichroism and dynamic multimode spectroscopy. Our results showed that cHSPA6 has similar binding affinity for both ATP and ADP (K D = ~50 nM). Binding of ATP and ADP reduced the surface hydrophobicity of the protein, and slightly altered its secondary structure, suggesting localized conformational rearrangement after ATP or ADP binding. Dynamic multimode spectroscopy revealed that cHSPA6 unfolds through three transitions with melting points (T m) of 42.3 ± 0.2, 61.3 ± 0.1, and 81.2 ± 0.2 °C. To the best of the author's knowledge, and literature search, this is the first report of the spectroscopic and thermodynamic properties of the Arabian camel heat shock protein.

Optimization of storage and stability of camel liver glutathione S-transferase.

Glutathione S-transferases (GSTs) are multifunctional enzymes and play an important role in cellular detoxification. Besides this, GSTs act as cytosolic carrier proteins that bind hydrophobic compounds such as heme, bilirubin, steroids, and polycyclic hydrocarbons. GST has great importance in biotechnology, as it is a target for vaccine and drug development and biosensors development for xenobiotics. Moreover, the GST tag has been extensively used for protein expression and purification. Until now, biophysical properties of camel liver GST have not been characterized. In the present study we have purified camel (Camelus dromedarius) liver GST to homogeneity in a single step by affinity chromatography with 23.4-fold purification and 60.6% yield. Our results showed that maximal activity of GST was at pH 6.5 and it was stable in the pH range of 5 to 10. The optimum temperature was 55°C and the Tm was 57°C. The chemical chaperone glycerol (3.3 M) was able to protect GST activity and aggregation against thermal denaturation by stabilizing the protein structure at 50 and 57°C, respectively. However, L-arginine (125 mM) did not protect GST against thermal stress. Far-ultraviolet circular dichroism (CD) spectra showed that glycerol protected the secondary structure of GST while L-arginine induced conformational changes under thermal stress. In conclusion, our studies on the GST stability suggest that glycerol works as a stabilizer and L-arginine acts as a destabilizer.

Cytosolic selection systems to study protein stability.

Here we describe biosensors that provide readouts for protein stability in the cytosolic compartment of prokaryotes. These biosensors consist of tripartite sandwich fusions that link the in vitro stability or aggregation susceptibility of guest proteins to the in vivo resistance of host cells to the antibiotics kanamycin, spectinomycin, and nourseothricin. These selectable markers confer antibiotic resistance in a wide range of hosts and are easily quantifiable. We show that mutations within guest proteins that affect their stability alter the antibiotic resistances of the cells expressing the biosensors in a manner that is related to the in vitro stabilities of the mutant guest proteins. In addition, we find that polyglutamine tracts of increasing length are associated with an increased tendency to form amyloids in vivo and, in our sandwich fusion system, with decreased resistance to aminoglycoside antibiotics. We demonstrate that our approach allows the in vivo analysis of protein stability in the cytosolic compartment without the need for prior structural and functional knowledge.

Benzophenone C-glucosides and gallotannins from mango tree stem bark with broad-spectrum anti-viral activity.

The high mutation rate of RNA viruses has resulted in limitation of vaccine effectiveness and increased emergence of drug-resistant viruses. New effective antivirals are therefore needed to control of the highly mutative RNA viruses. The n-butanol fraction of the stem bark of Mangifera indica exhibited inhibitory activity against influenza neuraminidase (NA) and coxsackie virus 3C protease. Bioassay guided phytochemical study of M. indica stem bark afforded two new compounds including one benzophenone C-glycoside (4) and one xanthone dimer (7), together with eleven known compounds. The structures of these isolated compounds were elucidated on the basis of spectroscopic evidences and correlated with known compounds. Anti-influenza and anti-coxsackie virus activities were evaluated by determining the inhibition of anti-influenza neuraminidase (NA) from pandemic A/RI/5+/1957 H2N2 influenza A virus and inhibition of coxsackie B3 virus 3C protease, respectively. The highest anti-influenza activity was observed for compounds 8 and 9 with IC50 values of 11.9 and 9.2µM, respectively. Compounds 8 and 9 were even more potent against coxsackie B3 virus 3C protease, with IC50 values of 1.1 and 2.0µM, respectively. Compounds 8 and 9 showed weak cytotoxic effect against human hepatocellular carcinoma and human epithelial carcinoma cell lines through MTT assay.

Probing the interaction mechanisms between sunset yellow dye and trypsin protein leading to amorphous aggregation under low pH conditions.

Protein aggregation poses a significant concern in the field of food sciences, and various factors, such as synthetic food dyes, can contribute to protein aggregation. One such dye, Sunset Yellow (SY), is commonly employed in the food industry. Trypsin was used as a model protein to assess the impact of SY. We employed several biophysical techniques to examine the binding and aggregation mechanisms between SY and trypsin at different pHs. Results from intrinsic fluorescence measurements indicate a stronger interaction between SY and trypsin at pH 2.0 compared to pH 6.0. Turbidity data reveal trypsin aggregation in the presence of 0.05-3.0 mM SY at pH 2.0, while no aggregation was observed at pH 6.0. Kinetic data demonstrate a rapid, lag-phase-free SY-induced aggregation of trypsin. Circular dichroism analysis reveals that trypsin adopts a secondary structure in the presence of SY at pH 6.0, whereas at pH 2.0, the secondary structure was nearly lost with increasing SY concentrations. Furthermore, turbidity and kinetics data suggest that trypsin aggregation depends on trypsin concentrations and pH. Our study highlights potential health risks associated with the consumption of SY, providing insights into its impact on human health and emphasizing the necessity for further research in this field.

Anionic surfactant causes dual conformational changes in insulin.

Amyloid fibrillation is a process by which proteins aggregate and form insoluble fibrils that are implicated in several neurodegenerative diseases. In n this study, we aimed to investigate the impact of the negatively charged detergent sodium dodecyl sulfate (SDS) on insulin amyloid fibrillation at pH 7.4 and 2.0, as SDS has been linked to the acceleration of amyloid fibrillation in vitro, but the underlying molecular mechanism is not fully understood. Our findings show that insulin forms amyloid-like aggregates in the presence of SDS at concentrations ranging from 0.05 to 1.8 mM at pH 2.0, while no aggregates were observed at SDS concentrations greater than 1.8 mM, and insulin remained soluble. However, at pH 7.4, insulin remained soluble regardless of the concentration of SDS. Interestingly, the aggregated insulin had a cross-ß sheet secondary structure, and when incubated with higher SDS concentrations, it gained more alpha-helix. The electrostatics and hydrophobic interaction of SDS and insulin may contribute to amyloid induction. Moreover, the SDS-induced aggregation was not affected by the presence of salts. Furthermore, as the concentration of SDS increased, the preformed insulin amyloid induced by SDS began to disintegrate. Overall, our study sheds light on the mechanism of surfactant-induced amyloid fibrillation in insulin protein.

Molecular mechanism of insulin aggregation in the presence of a cationic surfactant.

Protein aggregation and amyloid fibrillation are connected with neurodegenerative disorders. Insulin, a small molecular weight protein related to type II diabetes, has been shown to self-assemble to form protein aggregates. In this work, we investigated the effects of cetyltrimethylammonium bromide (CTAB) of insulin on the in vitro aggregation process at pH 7.4 and 2.0. The aggregation tendency of insulin was measured using a variety of biophysical approaches, including turbidity measurements, light scattering, far UV-CD, ThT dye binding, and transmission electron microscopy. The turbidity results demonstrated that at pH 7.4, a low concentration of CTAB (30-180 µM) causes insulin aggregation but at higer concentration (>180 µM) aggregation was not seen. However, at pH 2.0, both low as well as high concentrations of CTAB were unable to promote insulin aggregation. The ThT dye binding and far-UV CD data suggest that aggregation induced by CTAB is not having an ordered structure. Insulin treated with higher concentrations (>180 µM) of CTAB, the insulin gained a secondary structure. The possible cause of inducing aggregation in insulin is electrostatic and hydrophobic interaction because insulin contains a net negative charge at pH 7.4 and no aggregation at pH 2.0 due to electrostatic repulsion.

SDBS induces multiple catalase conformations in a dose-dependent manner.

Amyloid fibrils have been linked to several incurable diseases. They are long and thin fibrous proteins that self-assemble into fibrils. Small molecules can stimulate amyloid fibrillation, but the mechanism by which this happens is not well understood. This study examined how a negatively charged benzene ring containing surfactant, sodium dodecylbenzene sulphonate (SDBS), affects the fibrillation of bovine liver catalase (BLC). After SDBS treatment, BLC conformational changes were examined in vitro using turbidity, RLS kinetics, intrinsic fluorescence, ThT fluorescence, far-UV CD, and TEM. BLC in the native state was alpha-helical at pH 7.4, while it was converted to a random coil structure at pH 2.0. Far-UV CD and intrinsic fluorescence data showed that at concentrations <0.1 mM of SDBS, randomly coiled BLC assumed a native-like alpha-helical structure. However, between 0.1 and 1.0 mM SDBS, BLC was aggregated. ThT fluorescence and far-UV CD measurements showed the amyloid-like structures in the aggregated BLC. At higher SDBS concentrations (>1.0 mM) at pH 2.0, BLC again attains a native-like alpha-helical structure. It is essential for therapeutic purposes to clearly understand the process underlying surfactant- or lipid-induced fibrillation.

Elucidation of kinetic and structural properties of eye lens ζ-crystallin: an in vitro and in silico approach.

The Arabian Camelus dromedarius contains significant concentration of eye lens ζ-crystallin. This enzyme is also present in other life forms including humans, however in lower catalytic amounts. The recombinant camel ζ-crystallin was expressed in the E. coli BL21 (DE3) pLysS strain and purified using HisTrap column. The Km of the enzyme for 9,10-phenanthrenequinone (9,10-PQ) substrate and NADPH cofactor was determined to be 11.66 and 50.93 µM, respectively. The Vmax for 9,10-PQ and NADPH was obtained as 23.19 and 19.98 µM min-1, respectively. The optimum activity of the purified enzyme was found to be at pH 6.0 and at 55 °C. Different physico-chemical parameters were analysed including instability index (II), aliphatic index (AI) and the GRAVY index to establish proper characterization. The sequence of the recombinant ζ-crystallin was subjected to homology modelling using SWISS-MODEL webserver followed by validation of the modelled target structure. The evaluation of the modelled ζ-crystallin was performed by several parameters including Ramachandran plot, Z-score values followed by molecular dynamics (MD) simulation. The cumulative analysis of the physico-chemical, quantitative, qualitative and the essential dynamics of simulation of ζ-crystallin and its complexes with 9,10-PQ and NADPH helped in verifying the acceptable quality and stability of the ζ-crystallin structure.Communicated by Ramaswamy H. Sarma.

Bovine liver catalase turns into three conformational states after exposure to an anionic surfactant.

The mechanism by which anionic surfactants promote amyloid fibril is not well understood. Here, we investigated how sodium dodecyl sulfate (SDS), a negatively charged surfactant, affects the fibrillation of the partially unfolded random-coiled bovine liver catalase (BLC) at a pH of 2.0. We used several methods, including turbidity, RLS kinetics, intrinsic fluorescence, ThT fluorescence, far-UV CD, and TEM imaging, to evaluate the conformational changes of BLC in vitro in response to SDS treatment. BLC is a multimeric protein and well folded at physiological pH but forms a random coil structure at pH 2.0. Intrinsic fluorescence and far-UV CD data showed that below 0.1 mM SDS, random coiled BLC turned into a native-like structure. BLC incubated with an SDS concentration ranging from 0.1 to 2.0 mM led to the formation of aggregates. The ThT fluorescence intensity was enhanced in the aggregated BLC samples (0.1-2.0 mM SDS), and cross beta-sheeted structure was detected by the far UV CD measurements. BLC adopts a complete alpha-helical structure upon interacting with SDS at a more than 2.0 mM concentration at pH 2.0. Understanding the mechanism of surfactant- or lipid-induced fibrillation is important for therapeutic purposes.

SDS induces amorphous, amyloid-fibril, and alpha-helical structures in the myoglobin in a concentration-dependent manner.

Amyloid fibrils have been linked to a number of diseases. Surfactants imitate plasma membrane lipids and induce amyloid fibrils. This study examined the effects of the anionic surfactant sodium dodecyl sulfate (SDS) at pH 4.5 on equine skeletal muscle myoglobin (E-Mb). To analyze the effect of SDS on aggregation and amyloid-fibril formation to E-Mb, we used various spectroscopic techniques (turbidity, light scattering, intrinsic fluorescence, ThT fluorescence, and circular dichroism (CD)), electrophoretic, and microscopic techniques. Turbidity, SDS-PAGE, and light scattering all indicated the formation of E-Mb aggregates at SDS concentrations ranging from 0.2 mM to 1.0 mM. In the presence of 0.4 mM SDS, far-UV CD and TEM data indicate that E-MB forms amorphous aggregates. ThT binding, Far-UV CD, and TEM findings indicate that E-Mb forms amyloid-like structures in the presence of 0.6-1.0 mM SDS. However, no aggregation was seen at SDS concentrations above 1 mM. In the presence of high SDS concentrations (> 1 mM), the E-Mb exhibited native-like α-helical structure. As a result, SDS exhibited three distinct behaviors: amorphous aggregates, amyloid-fibrils, and helix-inducer. These findings also shed light on how amyloid fibrils are formed when anionic surfactants are introduced, which is a significant takeaway.

An insight into the binding and inhibition of eye ζ-crystallin by the environmental toxin arsenic: implications in eye diseases.

Arsenic contamination is highly prevalent in food chain, soil and groundwater. Continuous exposure to elevated levels of this environmental toxin is a global concern. Studies have reported enriched accumulation of arsenic in the eyes compared to other body organs leading to various eye diseases. Here, the impact of arsenic exposure on the enzymatic eye ζ-crystallin has been investigated. Arsenic inhibited the activity of the enzyme with an IC50 value of 35 µM. It decreased the free thiol group content of ζ-crystallin due to protein oxidation. The binding of arsenic with ζ-crystallin was explored using biophysical and computational tools. The enzyme undergoes some conformational changes upon arsenic binding. The binding constant (Kb) was determined to be 1.2 × 102 M-1. Thermodynamic parameters were determined by isothermal titration calorimetry (ITC) and the binding energy (ΔG) was calculated to be -3.52 kcal/mol. Molecular docking studies helped in visualizing the amino acid residues (especially Cys165) of the enzyme involved in binding with arsenic. Continuous arsenic exposure is expected to increase the eye crystallin-related abnormalities, elevating the risk of cataractogenesis. Therefore, proper measures need to be taken by authorities to control the contamination of arsenic in the environment and groundwater.Communicated by Ramaswamy H. Sarma.