Thiadiazole/Thiadiazine Derivatives as Insecticidal Agent: Design, Synthesis, and Biological Assessment of 1,3,4-(Thiadiazine/Thiadiazole)-Benzenesulfonamide Derivatives as IGRs Analogues against Spodoptera littoralis.

In keeping with our investigation, a simple and practical synthesis of novel heterocyclic compounds with a sulfamoyl moiety that can be employed as insecticidal agents was reported. The compound 2-hydrazinyl-N-(4-sulfamoylphenyl)-2-thioxoacetamide 1 was coupled smoothly with triethylorthoformate or a variety of halo compounds, namely phenacyl chloride, chloroacetyl chloride, chloroacetaldehyde, chloroacetone, 1,3-dichloropropane, 1,2-dichloroethane, ethyl chloroformate, 2,3-dichloro-1,4-naphthoquinone, and chloroanil respectively, which afforded the 1,3,4-thiadiazole and 1,3,4-thiadiazine derivatives. The new products structure was determined using elemental and spectral analysis. Under laboratory conditions, the biological and toxicological effects of the synthetic compounds were also evaluated as insecticides against Spodoptera littoralis (Boisd.). Compounds 3 and 5 had LC50 values of 6.42 and 6.90 mg/L, respectively. The investigated compounds (from 2 to 11) had been undergoing molecular docking investigation for prediction of the optimal arrangement and strength of binding between the ligand (herein, the investigated compounds (from 2 to 11)) and a receptor (herein, the 2CH5) molecule. The binding affinity within docking score (S, kcal/mol) ranged between -8.23 (for compound 5), -8.12 (for compound 3) and -8.03 (for compound 9) to -6.01 (for compound 8). These compounds were shown to have a variety of binding interactions within the 2CH5 active site, as evidenced by protein-ligand docking configurations. This study gives evidence that those compounds have 2CH5-inhibitory capabilities and hence may be used for 2CH5-targeting development. Furthermore, the three top-ranked compounds (5, 3, and 9) and the standard buprofezin were subjected to density functional theory (DFT) analysis. The highest occupied molecular orbital-lowest unoccupied molecular orbital (HOMO-LUMO) energy difference (ΔE) of compounds 5, 3, and 9 was found to be comparable to that of buprofezin. These findings highlighted the potential and relevance of charge transfer at the molecular level.

Identification, crystallization, and first X-ray structure analyses of phenyl boronic acid-based inhibitors of human carbonic anhydrase-II.

Human carbonic anhydrases (hCAs) play a central role in various physiological processes in the human body. HCAs catalyze the reversible hydration of CO2 into HCO3-, and hence maintains the fluid and pH balance. Overexpression of CA II is associated with diseases, such as glaucoma, and epilepsy. Therefore, CAs are important clinical targets and inhibition of different isoforms, especially hCA II is used in treatment of glaucoma, altitude sickness, and epilepsy. Therapeutically used CA inhibitors (CAI) are sulfonamide-based, such as acetazolamide, dichlorphenamide, methazolamide, ethoxzolamide, etc. However, they exhibit several undesirable effects such as numbness, tingling of extremities, malaise, metallic taste, fatigue, renal calculi, and metabolic acidosis. Therefore, there is an urgent need to identify safe and effective inhibitors of the hCAs. In this study, different phenyl boronic acids 1-5 were evaluated against bovine (bCA II) and hCA II. Among all, compound 1 (4-acetylphenyl boronic acid) was found to be active against bCAII and hCA II with IC50 values of 246 ± 0.48 and 281.40 ± 2.8 µM, respectively, while the remaining compounds were found in-active. Compound 1 was identified as competitive inhibitor of hCA II enzyme (Ki = 283.7 ± 0.002 µM). Additionally, compound 1 was found to be non-toxic against BJ Human fibroblast cell line. The X-ray crystal structure for hCA II in-complex with compound 1 was evaluated to a resolution of 2.6 Å. In fact, this the first structural analysis of a phenyl boron-based inhibitor bound to hCA II, allowing an additional structure-activity analysis of the compounds. Compound 1 was found to be directly bound in the active site of hCA II by interacting with His94, His119, and Thr199 residues. In addition, a bond of 3.11 Å between the zinc ion and coordinated boron atom of the boronic acid moiety of compound 1 was also observed, contributing to binding affinity of compound 1 for hCA II. PDB ID: 8IGF.

Overcoming aggregation with laser heated nanoelectrospray mass spectrometry: thermal stability and pathways for loss of bicarbonate from carbonic anhydrase II.

Variable temperature electrospray mass spectrometry is useful for multiplexed measurements of the thermal stabilities of biomolecules, but the ionization process can be disrupted by aggregation-prone proteins/complexes that have irreversible unfolding transitions. Resistively heating solutions containing a mixture of bovine carbonic anhydrase II (BCAII), a CO2 fixing enzyme involved in many biochemical pathways, and cytochrome c leads to complete loss of carbonic anhydrase signal and a significant reduction in cytochrome c signal above ∼72 °C due to aggregation. In contrast, when the tips of borosilicate glass nanoelectrospray emitters are heated with a laser, complete thermal denaturation curves for both proteins are obtained in <1 minute. The simultaneous measurements of the melting temperature of BCAII and BCAII bound to bicarbonate reveal that the bicarbonate stabilizes the folded form of this protein by ∼6.4 °C. Moreover, the temperature dependences of different bicarbonate loss pathways are obtained. Although protein analytes are directly heated by the laser for only 140 ms, heat conduction further up the emitter leads to a total analyte heating time of ∼41 s. Pulsed laser heating experiments could reduce this time to ∼0.5 s for protein aggregation that occurs on a faster time scale. Laser heating provides a powerful method for studying the detailed mechanisms of cofactor/ligand loss with increasing temperature and promises a new tool for studying the effect of ligands, drugs, growth conditions, buffer additives, or other treatments on the stabilities of aggregation-prone biomolecules.

1H, 15N and13C resonance assignments of S2A mutant of human carbonic anhydrase II.

In preparation for a detailed exploration of the structural and functional aspects of the Ser2Ala mutant of human carbonic anhydrase II, we present here almost complete sequence-specific resonance assignments for 1H, 15N, and 13C. The mutation of serine to alanine at position 2, located in the N-terminal region of the enzyme, significantly alters the hydrophilic nature of the site, rendering it hydrophobic. Consequently, there is an underlying assumption that this mutation would repel water from the site. However, intriguingly, comparative analysis of the mutant structure with the wild type reveals minimal discernible differences. These assignments serve as the basis for in-depth studies on histidine dynamics, protonation states, and its intricate role in protein-water interactions and catalysis.

Explainable artificial intelligence in the design of selective carbonic anhydrase I-II inhibitors via molecular fingerprinting.

Inhibiting the enzymes carbonic anhydrase I (CA I) and carbonic anhydrase II (CA II) presents a potential avenue for addressing nervous system ailments such as glaucoma and Alzheimer's disease. Our study explored harnessing explainable artificial intelligence (XAI) to unveil the molecular traits inherent in CA I and CA II inhibitors. The PubChem molecular fingerprints of these inhibitors, sourced from the ChEMBL database, were subjected to detailed XAI analysis. The study encompassed training 10 regression models using IC50 values, and their efficacy was gauged using metrics including R2, RMSE, and time taken. The Decision Tree Regressor algorithm emerged as the optimal performer (R2: 0.93, RMSE: 0.43, time-taken: 0.07). Furthermore, the PFI method unveiled key molecular features for CA I inhibitors, notably PubChemFP432 (C(O)N) and PubChemFP6978 (C(O)O). The SHAP analysis highlighted the significance of attributes like PubChemFP539 (C(O)NCC), PubChemFP601 (C(O)OCC), and PubChemFP432 (C(O)N) in CA I inhibitiotable n. Likewise, features for CA II inhibitors encompassed PubChemFP528(C(O)OCCN), PubChemFP791 (C(O)OCCC), PubChemFP696 (C(O)OCCCC), PubChemFP335 (C(O)NCCN), PubChemFP580 (C(O)NCCCN), and PubChemFP180 (C(O)NCCC), identified through SHAP analysis. The sulfonamide group (S), aromatic ring (A), and hydrogen bonding group (H) exert a substantial impact on CA I and CA II enzyme activities and IC50 values through the XAI approach. These insights into the CA I and CA II inhibitors are poised to guide future drug discovery efforts, serving as a beacon for innovative therapeutic interventions.

XFEL structure of carbonic anhydrase II: a comparative study of XFEL, NMR, X-ray and neutron structures.

The combination of X-ray free-electron lasers (XFELs) with serial femtosecond crystallography represents cutting-edge technology in structural biology, allowing the study of enzyme reactions and dynamics in real time through the generation of `molecular movies'. This technology combines short and precise high-energy X-ray exposure to a stream of protein microcrystals. Here, the XFEL structure of carbonic anhydrase II, a ubiquitous enzyme responsible for the interconversion of CO2 and bicarbonate, is reported, and is compared with previously reported NMR and synchrotron X-ray and neutron single-crystal structures.

Sulfonamides stimulate ROS formation upon glycation of human carbonic anhydrase II.

Advanced glycation end products are the most important species of glycation pathway, and cause disorders such as oxidative stress and diabetes. Sulfonamide compounds, which are generally known as widespread inhibitors, are potential agents used in different drug products, which can readily enter biological matrices. In the present work, the structure and activity of human carbonic anhydrase II studied in the presence of glucose as well as four sulfonamide agents from different views. These included enzyme kinetics, free lysine content, fluorescence spectroscopy, circular dichroism, and ROS measurement. Our results indicated that upon glycation, the structure of HCA II collapses and 8 to 13 lysine residues will be more available based on ligand incubation. Secondary and tertiary structural changes were also observed in the presence and absence of sulfonamide agents using fluorescence and circular dichroism methods, respectively. These spectroscopic data also showed a remarkable increase in hydrophobicity and decrease in α-helix contents during glycation, especially after 35 days of incubation. ROS assay was studied in the presence of glucose and sulfonamide compounds, that demonstrated the role of sulfonamide compounds in ROS formation in the presence of glucose in a synergistic manner. Overall, our data indicated that sulfonamides act as a stimulant factor upon prolonged interaction with HCA II and may intensify the complications of some disorders, such as diabetes and other conformational diseases.

pH-Dependent Structure and Dynamics of the Catalytic Domains of Human Carbonic Anhydrase II and IX.

Extensive computer simulation studies have been carried out to probe the pH-dependent structure and dynamics of the two most efficient isoenzymes II and IX of human carbonic anhydrase (HCA) that control the pH in the human body. The equilibrium structure and hydration of their catalytic domains are found to be largely unaffected by the variation of pH in the range studied, in close agreement with the known experimental results. In contrast, a significant effect of the change in pH is observed for the first time on the local electrostatic potential of the active site walls and the dynamics of active site water molecules. We also report for the first time the free energy and kinetics of coupled fluctuations of orientation and protonation states of the well-known His-mediated proton shuttle (His-64) in both isozymes at pH 7 and 8. The transitions between different tautomers of in or out conformations of His-64 side chain range between 109 and 106 s-1 depending on pH. Possible implications of these results on conformation-dependent pKa of His-64 side chain and its role in driving the catalysis toward hydration of CO2 or dehydration of HCO3- with varying pH are discussed.

Investigation of the effects of some pesticides on carbonic anhydrase isoenzymes.

The aim of this study was to investigate the inhibitory effects of some pesticides known to have harmful effects on human health on carbonic anhydrase isoenzymes. Therefore, carbonic anhydrase isoenzymes (hCA I and II) were purified from human erythrocytes. The isoenzymes were purified from human erythrocytes by using an affinity column that has the chemical structure of Sepharose-4B-4-(6-amino-hexyloxy)-benzenesulfonamide. The purity of the isoenzymes was checked by sodium dodecyl sulphate polyacrylamide gel electrophoresis (SDSPAGE). It was determined that the pesticides used in this study inhibit hCA I and hCA II isoenzymes at different levels in vitro. It was determined that the strongest inhibitor for the hCA I enzyme was Carbofuran (IC50 :6.52 µM; Ki : 3.58 µM) and the weakest one was 1-Naphtol (IC50 :16.55 µM; Ki : 14.4 µM) among these pesticides. It was also found that the strongest inhibitor for the hCA II enzyme was coumatetralil (IC50 :5.06 µM; Ki : 1.62 µM) and the weakest one was Dimethachlor (IC50 14.6 µM; Ki : 8.44 µM).

A Case Study on the Use of Binding Free Energies to Screen Inhibitors of Human Carbonic Anhydrase II.

We present in this article a case study on the thermodynamics of binding to human carbonic anhydrase II (HCA II) by three well-known inhibitors, viz. (a) acetazolamide (AZM) that directly binds to the catalytic Zn(II) ion at the active site, (b) non-zinc binding 6-hydroxy-2-thioxocoumarin (FC5) (c) 2-[(S)-benzylsulfinyl]benzoic acid (3G1). In each case, the crystal structure or its analogue of inhibitor-bound HCA II has been used to perform classical molecular dynamics (MD) simulation in water till 1 µ s ${1\hskip0.33em\mu s}$ . AZM and FC5 are found to undergo repeated binding and unbinding with markedly different dynamics from the partially buried, substrate-binding hydrophobic pocket near the active site. 3G1, on the other hand, is found to remain mostly at its crystallographic binding site occluded from the active site of HCA II. The associated binding free energies ( Δ G b i n d , s o l v ${{\rm \Delta }{G}_{bind,solv}}$ ) have been computed using the known MM/GBSA method and compared to the available experimental data. Our results show that Δ G b i n d , s o l v ${{\rm \Delta }{G}_{bind,solv}}$ encounters several issues including limited sampling of multiple binding sites and incorrect prediction of the affinity of the chosen ligands. Possible use of the simulation results in further construction of Markov state models is also discussed.

A Series of Trifluoromethylisoxazolyl- and Trifluoromethylpyrazolyl- Substituted (Hetero)aromatic Sulfonamide Carbonic Anhydrase Inhibitors: Synthesis, and Convenient Prioritization Workflow for Further In Vivo Studies.

AIMS: To synthesize novel sulfonamide inhibitors of carbonic anhydrase and develop in vitro prioritization workflow to select compounds for in vivo evaluation. BACKGROUND: Carbonic anhydrase (CA) inhibitors gain significant attention in the context of drug discovery research for glaucoma, hypoxic malignancies, and bacterial infections. In previous works, we have successfully used direct sulfochlorination approach to develop diverse heterocyclic primary sulfonamides with remarkable activity and selectivity against therapeutically relevant CA isoforms. OBJECTIVE: Synthesis and investigation of the CA inhibitory properties of novel trifluoromethylisoxazolyl- and trifluoromethylpyrazolyl-substituted (hetero)aromatic sulfonamides. METHODS: Thirteen trifluoromethylisoxazolyl- and thirteen trifluoromethylpyrazolyl-substituted (hetero) aromatic sulfonamides were synthesized by direct sulfochlorination of hydroxyisoxazolines and pyrazoles followed by reaction with ammonia. The compound structures were confirmed by 1H and 13C NMR as well as element analysis. The obtained compounds were evaluated, using the CA esterase activity assay, for their potential to block the catalytic activity of bovine CA (bCA). RESULTS: Eight most potent compounds selected based on the esterase activity assay data were tested for direct affinity to the enzyme using the thermal shift assay (TSA). These compounds displayed Kd values (measured by TSA) in the double-digit nanomolar range, thus showing comparable activity to the reference drug acetazolamide. CONCLUSION: Coupling the bCA esterase activity assay with thermal shift assay represents a streamlined and economical strategy for the prioritization of sulfonamide CA inhibitors for subsequent evaluation in vivo.

Acetylphenyl-substituted imidazolium salts: synthesis, characterization, in silico studies and inhibitory properties against some metabolic enzymes.

Herein, we present how to synthesize thirteen new 1-(4-acetylphenyl)-3-alkylimidazolium salts by reacting 4-(1-H-imidazol-1-yl)acetophenone with a variety of benzyl halides that contain either electron-donating or electron-withdrawing groups. The structures of the new imidazolium salts were conformed using different spectroscopic methods (1H NMR, 13C NMR, 19F NMR, and FTIR) and elemental analysis techniques. Furthermore, these compounds' the carbonic anhydrase (hCAs) and acetylcholinesterase (AChE) enzyme inhibition activities were investigated. They showed a highly potent inhibition effect toward AChE and hCAs with Ki values in the range of 8.30 ± 1.71 to 120.77 ± 8.61 nM for AChE, 16.97 ± 2.04 to 84.45 ± 13.78 nM for hCA I, and 14.09 ± 2.99 to 69.33 ± 17.35 nM for hCA II, respectively. Most of the synthesized imidazolium salts appeared to be more potent than the standard inhibitor of tacrine (TAC) against AChE and Acetazolamide (AZA) against CA. In the meantime, to prospect for potential synthesized imidazolium salt inhibitor(s) against AChE and hCAs, molecular docking and an ADMET-based approach were exerted.

Synthesis, Biological Evaluation, and Molecular Dynamics of Carbothioamides Derivatives as Carbonic Anhydrase II and 15-Lipoxygenase Inhibitors.

A series of hydrazine-1-carbothioamides derivatives (3a-3j) were synthesized and analyzed for inhibitory potential towards bovine carbonic anhydrase II (b-CA II) and 15-lipoxygenase (15-LOX). Interestingly, four derivatives, 3b, 3d, 3g, and 3j, were found to be selective inhibitors of CA II, while other derivatives exhibited CA II and 15-LOX inhibition. In silico studies of the most potent inhibitors of both b-CA II and 15-LOX were carried out to find the possible binding mode of compounds in their active site. Furthermore, MD simulation results confirmed that these ligands are stably bound to the two targets, while the binding energy further confirmed the inhibitory effects of the 3h compound. As these compounds may have a role in particular diseases, the reported compounds are of great relevance for future applications in the field of medicinal chemistry.

Ibuprofen: a weak inhibitor of carbonic anhydrase II.

Carbonic anhydrases (CAs) are drug targets for a variety of diseases. While many clinically relevant CA inhibitors are sulfonamide-based, novel CA inhibitors are being developed that incorporate alternative zinc-binding groups, such as carboxylic acid moieties, to develop CA isoform-specific inhibitors. Here, the X-ray crystal structure of human CA II (hCA II) in complex with the carboxylic acid ibuprofen [2-(4-isobutylphenyl)propanoic acid, a common over-the-counter nonsteroidal anti-inflammatory drug] is reported to 1.54 Šresolution. The binding of ibuprofen is overlaid with the structures of other carboxylic acids in complex with hCA II to compare their inhibition mechanisms by direct or indirect (via a water) binding to the active-site zinc. Additionally, enzyme-inhibition assays using ibuprofen, nicotinic acid and ferulic acid were performed with hCA II to determine their IC50 values and were compared with those of other carboxylic acid binders. This study discusses the potential development of CA inhibitors utilizing the carboxylic acid moiety.

Intrinsic affinity of protein - ligand binding by differential scanning calorimetry.

Differential scanning calorimetry (DSC) determines the enthalpy change upon protein unfolding and the melting temperature of the protein. Performing DSC of a protein in the presence of increasing concentrations of specifically-binding ligand yields a series of curves that can be fit to obtain the protein-ligand dissociation constant as done in the fluorescence-based thermal shift assay (FTSA, ThermoFluor, DSF). The enthalpy of unfolding, as directly determined by DSC, helps improving the precision of the fit. If the ligand binding is linked to protonation reactions, the intrinsic binding constant can be determined by performing the affinity determination at a series of pH values. Here, the intrinsic, pH-independent, affinity of acetazolamide binding to carbonic anhydrase (CA) II was determined. A series of high-affinity ligands binding to CAIX, an anticancer drug target, and CAII showed recognition and selectivity for the anticancer isozyme. Performing the DSC experiment in buffers of highly different enthalpies of protonation enabled to observe the ligand unbinding-linked protonation reactions and estimate the intrinsic enthalpy of binding. The heat capacity of combined unfolding and unbinding was determined by varying the ligand concentrations. Taken together, these parameters provided a detailed thermodynamic picture of the linked ligand binding and protein unfolding process.

Design, Synthesis and Molecular Docking Study of Novel 3-Phenyl-ß-Alanine-Based Oxadiazole Analogues as Potent Carbonic Anhydrase II Inhibitors.

Carbonic anhydrase-II (CA-II) is strongly related with gastric, glaucoma, tumors, malignant brain, renal and pancreatic carcinomas and is mainly involved in the regulation of the bicarbonate concentration in the eyes. With an aim to develop novel heterocyclic hybrids as potent enzyme inhibitors, we synthesized a series of twelve novel 3-phenyl-ß-alanine 1,3,4-oxadiazole hybrids (4a-l), characterized by 1H- and 13C-NMR with the support of HRESIMS, and evaluated for their inhibitory activity against CA-II. The CA-II inhibition results clearly indicated that the 3-phenyl-ß-alanine 1,3,4-oxadiazole derivatives 4a-l exhibited selective inhibition against CA-II. All the compounds (except 4d) exhibited good to moderate CA-II inhibitory activities with IC50 value in range of 12.1 to 53.6 µM. Among all the compounds, 4a (12.1 ± 0.86 µM), 4c (13.8 ± 0.64 µM), 4b (19.1 ± 0.88 µM) and 4h (20.7 ± 1.13 µM) are the most active hybrids against carbonic CA-II. Moreover, molecular docking was performed to understand the putative binding mode of the active compounds. The docking results indicates that these compounds block the biological activity of CA-II by nicely fitting at the entrance of the active site of CA-II. These compounds specifically mediating hydrogen bonding with Thr199, Thr200, Gln92 of CA-II.

Targeting Colorectal Cancer Cells with a Functionalised Calix[4]arene Receptor: Biophysical Studies.

Colorectal cancer (CRC) is a disease which is causing a high degree of mortality around the world. The present study reports the antiproliferative impact of the thioacetamide calix[4]arene, CAII receptor on a highly differentiated Caco-2 cell line. This statement is corroborated by the MTT assay results which revealed a reduction in the cell viability with an IC50 value of 19.02 ± 0.04 µM. Microscopic results indicated that at the starting amount of 10 µM of CAII, a decrease in cells confluency can already be observed in addition to changes in cells morphology. Cell metabolic pathway changes were also investigated. 1H NMR findings showed downregulation in lactate, pyruvate, phosphocholine, lipids, and hydroxybutyrate with the upregulation of succinate, indicating a decline in the cells proliferation. Some biochemical alterations in the cells as a result of the CAII treatment were found by Raman spectroscopy.

Synthesis, carbonic anhydrase enzyme inhibition evaluations, and anticancer studies of sulfonamide based thiadiazole derivatives.

The sulfonamide-based thiadiazole derivatives (STDs) with different hydrophobic/hydrophilic substitutions were synthesized to investigate their potentials in carbonic anhydrase inhibition (CAI). The CAI activity of the STDs (4a-4h) and the mechanism of the inhibition kinetics were determined. STD 4f contained both methoxy and Cl groups at benzene ring in STD 4f showed the lowest IC50 value. The molecular docking study confirmed that STDs bind strongly with the active sites of the target protein PDBID 1V9E. With the help of Lineweaver-Burk plots, inhibition kinetics of PDBIR 1V9E protein with STDs were determined. Cytotoxicity was checked against human keratinocyte cell lines and the anticancer properties were determined against MCF-7 cell lines. The electrochemical method was used to investigate the binding study with DNA and CA enzymes. Anticancer studies showed that STDs have weak bonding ability to DNA and strong binding ability with CA. It is concluded that anticancer activity is through CAI rather than by DNA binding.

Role of Thr199 residue in human ß-carbonic anhydrase-II pH-dependent activity elucidated by microsecond simulation analysis.

Carbonic anhydrases catalyze the reversible hydration of carbon dioxide to form bicarbonate, a reaction required for many functions such as carbon assimilation, pH acid-base homeostasis, respiration and photosynthesis via a zinc-hydroxide mechanism for carbon dioxide hydration. In earlier studies, it was revealed that Carbonic anhydrases are inactive at pH 7.5 and active at pH 8.4. This steep pH dependence for its activity led us to design this work to understand its mode of action at atomic level detail. In our microsecond simulation based analysis, it was revealed that the interaction between Glu106 and Thr199 plays a critically important role in its activity. Thr199 co-ordinated loop movement was observed to be acting as a lid, with 'open' and 'close' mechanism for substrate entry to the core of the catalytic site, where Zn-ion resides and executes its carbon dioxide hydration mechanism. On the other hand, decline in the total secondary structural elements percentage in the protein was observed in correspondence to the pH condition change. The α-helices between Thr125-Gly145 and Val150-Lys170 residues were especially noticed to be losing their structural integrity responsible for formation of dimer and tetramers. In conclusion, our analysis showed that the interaction between Glu106 and Thr199 is crucial for maintaining the structural integrity of the Thr199 coordinated loop, responsible for allowing substrate towards the catalytic site.Communicated by Ramaswamy H. Sarma.

Microsecond simulation analysis of carbonic anhydrase - II in complex with (+)-cathechin revealed molecular interactions responsible for its amelioration effect on fluoride toxicity.

Fluorosis is a chronic condition caused by overexposure to fluoride, marked by impaired dental, skeletal, and non-skeletal health. In presence of excess fluoride ions, in severe cases calcification of the ligaments observed. Earlier studies have suggested that the disruption of carbonic anhydrase activity via ionic homeostasis change was associated with F toxicity. In a recent study, it was demonstrated that Tamarind fruit extract was effective in increasing the urinary F excretion in male Wistar rats via studying the mRNA expression of carbonic anhydrase II (CA II) in kidney homogenates using western blotting, immunohistochemistry and quantitative Realtime PCR based studies. We have carried out this study to understand the detailed molecular level interactions responsible for this tamarind extract based (+)-cathechin compound towards lowering the F toxicity via targeting CA-II. From our study, it was revealed that due to the ability of (+)-cathechin compound to bind tightly filling complete available space at the catalytically important site forming metal coordinated ionic bonds with His94, His96 and His119 residues helps in restricting F ions to interact with Zn ion located at the core of catalytic site responsible for its functionality. On the other hand, interaction of (+)-cathechin compound with Gln92 was observed to be critically important towards inducing conformational changes in CA-II, thus allowing (+)-cathechin compound to burry even deeply inside the catalytic site.Communicated by Ramaswamy H. Sarma.