Binding Affinity Of Enzyme Research Articles

Synthesis and Biological Evaluation of 1,2,3-Triazole Appended Benzothiazinone Derivatives via Click Chemistry

: We have created novel 1,2,3-triazole-based benzothiazinone derivatives in the current work. The produced compounds' in vitro antioxidant, antifungal, and antitubercular properties were assessed. Moreover, a simulated molecular docking analysis was conducted on the cytochrome P450 lanosterol 14α-demethylase active site to elucidate the enzyme's binding affinity and interactions with synthesised benzothiazinone derivatives. A notable correlation between these compounds' antifungal activity and binding score was indicated by molecular docking data. The synthetic 1,2,3-triazole-based benzothiazinone derivatives may satisfy the structural criteria for creating novel antifungal drugs, according to the results of the in vitro and in silico investigations.

Allosteric mechanism of synergistic effect in α- and β-amylase mixtures

Alpha-amylase and beta-amylase coexist as mixtures in industrial production, and the two amylases have active synergistic effects when they approach each other. These effects are due to enhanced enzyme binding affinity for the substrate and the rate of particle hydrolysis. Here, we report the allosteric mechanism of this synergistic effect in α- and β-amylase mixtures. The assay showed higher activity after mixing α- and β-amylase. Molecular docking showed that α- and β-amylase create a stable dual-enzyme complex with high binding energy, and that complex formation does not affect the exposure of respective active sites. β-Amylase is specifically bound to the B domain of α-amylase, and the dynamic plasticity of the B domain makes it move spatially, and this adjustment leads to a more open conformation in the active site of α-amylase. Because the enzymes binding make the complex more stable, the degree to which the relative activity of the dual-enzyme complex is inhibited is significantly reduced. After enzyme hydrolysis, the products maltose and glucose accumulate and produce competitive inhibition, which explains the relative activity decrease of the later-stage dual-enzyme cooperation. Structural characterization by FT-IR and CD spectroscopy did not reveal significant changes in respective secondary structures after enzyme binding.

Insights into the Effects of Ligand Binding on Leucyl-tRNA Synthetase Inhibitors for Tuberculosis: In Silico Analysis and Isothermal Titration Calorimetry Validation.

Incidences of drug-resistant tuberculosis have become common and are rising at an alarming rate. Aminoacyl t-RNA synthetase has been validated as a newer target against Mycobacterium tuberculosis. Leucyl t-RNA synthetase (LeuRS) is ubiquitously found in all organisms and regulates transcription, protein synthesis, mitochondrial RNA cleavage, and proofreading of matured t-RNA. Leucyl t-RNA synthetase promotes growth and development and is the key enzyme needed for biofilm formation in Mycobacterium. Inhibition of this enzyme could restrict the growth and development of the mycobacterial population. A database consisting of 2734 drug-like molecules was screened against leucyl t-RNA synthetase enzymes through virtual screening. Based on the docking scores and MMGBSA energy values, the top three compounds were selected for molecular dynamics simulation. The druggable nature of the top three hits was confirmed by predicting their pharmacokinetic parameters. The top three hits-compounds 1035 (ZINC000001543916), 1054 (ZINC000001554197), and 2077 (ZINC000008214483)-were evaluated for their binding affinity toward leucyl t-RNA synthetase by an isothermal titration calorimetry study. The inhibitory activity of these compounds was tested against antimycobacterial activity, biofilm formation, and LeuRS gene expression potential. Compound 1054 (Macimorelin) was found to be the most potent molecule, with better antimycobacterial activity, enzyme binding affinity, and significant inhibition of biofilm formation, as well as inhibition of the LeuRS gene expression. Compound 1054, the top hit compound, has the potential to be used as a lead to develop successful leucyl t-RNA synthetase inhibitors.

Utilizing noncatalytic ACE2 protein mutant as a competitive inhibitor to treat SARS-CoV-2 infection.

Angiotensin converting-enzyme 2 (ACE2) is an enzyme catalyzing the conversion of angiotensin 2 into angiotensin 1-7. ACE2 also serves as the receptor of several coronaviruses, including SARS-CoV-1 and SARS-CoV-2. Therefore, ACE2 could be utilized as a therapeutic target for treating these coronaviruses, ideally lacking enzymatic function. Based on structural analysis, specific mutations were introduced to generate mutants of ACE2 and ACE2-Fc (fusion protein of ACE2 and Fc region of IgG1). The enzyme activity, binding affinity, and neutralization abilities were measured. As predicted, five mutants (AMI081, AMI082, AMI083, AMI084, AMI090) have completely depleted ACE2 enzymatic activities. More importantly, enzyme-linked receptor-ligand assay (ELRLA) and surface plasmon resonance (SPR) results showed that 2 mutants (AMI082, AMI090) maintained binding activity to the viral spike proteins of SARS-CoV-1 and SARS-CoV-2. In An in vitro neutralization experiment using a pseudovirus, SARS-CoV-2 S1 spike protein-packed lentivirus particles, was also performed, showing that AMI082 and AMI090 significantly reduced GFP transgene expression. Further, in vitro virulent neutralization assays using SARS-CoV-2 (strain name: USA-WA1/2020) showed that AMI082 and AMI090 had remarkable inhibitory effects, indicated by comparable IC50 to wildtype ACE2 (5.33 µg/mL). In addition to the direct administration of mutant proteins, an alternative strategy for treating COVID-19 is through AAV delivery to achieve long-lasting effects. Therefore, AAV5 encoding AMI082 and AMI090 were packaged and transgene expression was assessed. In summary, these ACE2 mutants represent a novel approach to prevent or treat COVID-19 and other viruses with the same spike protein.

A highly susceptible hACE2-transgenic mouse model for SARS-CoV-2 research.

Several animal models have been used to assist the development of vaccines and therapeutics since the COVID-19 outbreak. Due to the lack of binding affinity of mouse angiotensin-converting enzyme II (ACE2) to the S protein of severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2), increasing the susceptibility of mice to SARS-CoV-2 infection was considered in several ways. Here, we generated a COVID-19 mouse model expressing human ACE2 (hACE2) under the control of the CAG promoter. Overexpression of hACE2 did not pose a significant effect on weight growth. After SARS-CoV-2 inoculation, mice showed obvious viral replication and production of inflammation within 7 days, with a gradual decrease in body weight until death. Virological testing found that the virus can replicate in the respiratory system, small intestine, and brain. Additionally, this mouse model was applied to compare two antibody drug candidates, the anti-RBD antibody (MW06) and the mouse CD24-conjugated anti-RBD antibody (mCD24-MW06). Differences in antiviral effects between these two antibodies can be demonstrated in this mouse model when a challenge dose that invalidates the anti-RBD antibody treatment was used. This study provided a new mouse model for studying SARS-CoV-2 pathogenesis and evaluating potential interventions.

Structural insights into binding of polyglutamylated tetrahydrofolate by serine hydroxymethyltransferase 8 from soybean.

Tetrahydrofolate and its derivatives participate in one-carbon transfer reactions in all organisms. The cellular form of tetrahydrofolate (THF) is modified by multiple glutamate residues and polyglutamylation plays a key role in organellar and cellular folate homeostasis. In addition, polyglutamylation of THF is known to increase the binding affinity to enzymes in the folate cycle, many of which can utilize polyglutamylated THF as a substrate. Here, we use X-ray crystallography to provide a high-resolution view of interactions between the enzyme serine hydroxymethyltransferase (SHMT), which provides one carbon precursors for the folate cycle, and a polyglutamylated form of THF. Our 1.7 Å crystal structure of soybean SHMT8 in complex with diglutamylated 5-formyl-THF reveals, for the first time, a structural rearrangement of a loop at the entrance to the folate binding site accompanied by the formation of novel specific interactions between the enzyme and the diglutamyl tail of the ligand. Biochemical assays show that additional glutamate moieties on the folate ligand increase both enzyme stability and binding affinity. Together these studies provide new information on SHMT structure and function and inform the design of anti-folate agents.

Uncovering the role of leucine 59 in Renilla luciferase stability and activity with error-prone PCR: Implications for protein engineering

A new variant of Renilla luciferase, named Met C-SRLuc 8, was obtained from a random mutagenesis library and expressed in Escherichia coli BL21 (DE3) plys and purified. The results of the enzyme's binding affinity, kinetic stability, and bioinformatic studies demonstrated that leucine 59, located within the hot-spot foldon in the N-terminal domain of the protein, plays a significant role in the stability and activity of Renilla luciferase. These findings may facilitate the engineering of different variants of this enzyme to achieve thermally stable versions for various biotechnological applications.

Expression and characterization of a novel PPO gene from Mucuna pruriens (L.) DC. var. pruriens involved in catecholamine pathway mediated synthesis of L-DOPA

Mucuna pruriens (L.) DC var. pruriens is a well-known legume of Fabaceae family. The plant is the natural source of l-DOPA, precursor molecule of dopamine neurotransmitter used in the treatment of Parkinson´s disease. In M. pruriens, the Catecholamine (CA) biosynthesis is mediated by Polyphenol oxidase (PPO) as validated in our previous reports. In the present investigation, we identified three PPO isoforms and were quantitatively analyzed by quantitative real-time PCR to understand the expression of PPO´s constitutively in the different tissues of M. pruriens. Consequently, the highly expressed PPO isoform was cloned, and the recombinant protein purified followed by functional characterization. The Kinetic analysis using Lineweaver-Burk curves showed greater binding affinity of enzyme towards its specific substrate. This study provides new insight into plant derived recombinant PPO enzyme, which is involved in CA biosynthetic pathway in this taxon. The Metabolic pathway engineering of PPO step would result in modulation of l-DOPA synthesis in this plant, in future.

Forrestiacids E-K: Further [4 + 2]-Type Triterpene-Diterpene Hybrids as Potential ACL Inhibitors from the Vulnerable Conifer Pseudotsuga forrestii.

Seven [4 + 2]-type triterpene-diterpene hybrids derived from a rearranged or a normal lanostane unit (dienophile) and an abietane moiety (diene), forrestiacids E-K (1-7, respectively), were further isolated and characterized from Pseudotsuga forrestii (a vulnerable conifer endemic to China). The intriguing molecules were revealed with the guidance of an LC-MS/MS-based molecular ion networking strategy combined with conventional phytochemical procedures. Their chemical structures with absolute configurations were established by spectroscopic data, chemical transformation, electronic circular dichroism calculations, and single-crystal X-ray diffraction analysis. They all contain a rare bicyclo[2.2.2]octene motif. Both forrestiacids J (6) and K (7) represent the first examples of this unique class of [4 + 2]-type hybrids that arose from a normal lanostane-type dienophile. Some isolates remarkably inhibited ATP-citrate lyase (ACL), with IC50 values ranging from 1.8 to 11 μM. Docking studies corroborated the findings by highlighting the interactions between the bioactive compounds and the ACL enzyme (binding affinities: -9.9 to -10.7 kcal/mol). The above findings reveal the important role of protecting plant species diversity in support of chemical diversity and potential sources of new therapeutics.

Squamabietenols A–F, undescribed abietane-O-abietane dimeric diterpenoids from the ornamental conifer Juniperus squamata and their ATP-citrate lyase inhibitory activities

Six undescribed naturally occurring abietane-O-abietane dimers (squamabietenols A–F) together with one 3,4-seco-totarane-type, a pimarane-type, and 17 related known mono-/dimeric diterpenoids were isolated and characterized from the needles and twigs of the ornamental conifer Juniperus squamata. The undescribed structures and their absolute configurations were established by extensive spectroscopic methods, GIAO NMR calculations with DP4+ probability analyses, and ECD calculations. Squamabietenols A and B showed significant inhibitory effects against ATP-citrate lyase (ACL, a novel drug target for hyperlipidemia and other metabolic disorders), with IC50 values of 8.82 and 4.49 μM, respectively. A molecular docking study corroborated the findings by highlighting the interactions between the bioactive compounds and the ACL enzyme (binding affinities: −7.1 to −9.0 kcal/mol). The unique abietane-O-abietane dimeric diterpenoids are quite rare in the vegetable kingdom, and they are of chemotaxonomic significance for the Cupressaceae family.

In Silico Identification of Lead Compounds for Pseudomonas Aeruginosa PqsA Enzyme: Computational Study to Block Biofilm Formation.

Pseudomonas aeruginosa is an opportunistic Gram-negative bacterium implicated in acute and chronic nosocomial infections and a leading cause of patient mortality. Pseudomonas aeruginosa infections are frequently associated with the development of biofilms, which give the bacteria additional drug resistance and increase their virulence. The goal of this study was to find strong compounds that block the Anthranilate-CoA ligase enzyme made by the pqsA gene. This would stop the P. aeruginosa quorum signaling system. This enzyme plays a crucial role in the pathogenicity of P. aeruginosa by producing autoinducers for cell-to-cell communication that lead to the production of biofilms. Pharmacophore-based virtual screening was carried out utilizing a library of commercially accessible enzyme inhibitors. The most promising hits obtained during virtual screening were put through molecular docking with the help of MOE. The virtual screening yielded 7/160 and 10/249 hits (ZINC and Chembridge). Finally, 2/7 ZINC hits and 2/10 ChemBridge hits were selected as potent lead compounds employing diverse scaffolds due to their high pqsA enzyme binding affinity. The results of the pharmacophore-based virtual screening were subsequently verified using a molecular dynamic simulation-based study (MDS). Using MDS and post-MDS, the stability of the complexes was evaluated. The most promising lead compounds exhibited a high binding affinity towards protein-binding pocket and interacted with the catalytic dyad. At least one of the scaffolds selected will possibly prove useful for future research. However, further scientific confirmation in the form of preclinical and clinical research is required before implementation.

Distinct roles of carbohydrate-binding modules in multidomain β-1,3-1,4-glucanase on polysaccharide degradation.

Lam16A is a novel GH16 β-1,3-1,4-lichenase isolated from the genus Caldicellulosiruptor which can utilize untreated carbohydrate components of plant cell walls. Its catalytic module has been characterized that the six carbohydrate-binding modules (CBMs) were queued in the C-terminus, but their roles were still unclear. Here, full-length and CBM-truncated mutants of Lam16A were purified and characterized through heterologous expression in Escherichia coli. The profiles of these proteins, including the enzyme activity, degrading efficiency, substrate-binding affinity, and thermostability, were explored. Full-length Lam16A with six CBMs showed excellent thermostability and the highest activity against barley β-glucan and laminarin with optimum pH of 6.5. The CBMs stimulated degrading ability of the catalytic module, especially against β-1,3(4)-glucan-based polysaccharides. The released products from β-1,3-1,4-glucan by Lam16A or its truncated mutants revealed an endo-type glycoside hydrolase. Lam16As exhibited strong binding affinities to the insoluble polysaccharides, especially Lam16A-1CBM. The degradation of yeast cell walls by Lam16A enzyme solution relative to the control reduced the absorbance values at OD800 by ~ 85% ± 1.2, enabling the release of up to ~ 0.057 ± 0.0039µg/mL of the cytoplasmic protein into the supernatant, lowering the viability of the cells by ~ 70.3% ± 6.9, thus causing significant damage in the cell wall structure. Taken together, CBMs could influence the substrate specificity, thermal stability, and binding affinity of β-1,3-1,4-glucanase. These results demonstrate the great potential of these enzymes to promote the bioavailability of β-1,3-glucan oligosaccharides for health benefits. KEY POINTS: • Carbohydrate-binding modules strongly influenced the enzyme activity and binding affinity, and further impacted glycoside hydrolase activity. • Lam16A enzymes have sufficient ability to hydrolyze β-1,3-1,4-glucan-based polysaccharides. • Lam16As provide a powerful tool to promote the bioavailability of β-1,3-glucan oligosaccharides.

Synthesis of substituted norcaranes for use as probes of enzyme mechanisms.

Norcarane is a mechanistic probe of monooxygenase enzymes that is able to detect the presence of cationic or radical intermediates. The addition of substituents around the bicycloheptane ring of the norcarane scaffold can assist in improving enzyme binding affinity and thus improve the regioselectivity of oxidation. Here we prepare in three-step, diastereoselective syntheses, ten norcaranes monosubstituted α to the cyclopropane as advanced probes. Four of these compounds were examined in enzyme binding experiments to evaluate their potential as probe substrates. Additionally, 19 potential products of enzymatic oxidation were generated via two additional synthetic steps for use as product standards in future studies.

In silico study of SARS-CoV-2 spike protein RBD and human ACE-2 affinity dynamics across variants and Omicron subvariants.

The coronavirus disease 2019virus outbreak continues worldwide, with many variants emerging, some of which are considered variants of concern (VOCs). The WHO designated Omicron as a VOC and assigned it under variant B.1.1.529. Here, we used computational studies to examine the VOCs, including Omicron subvariants,and one variant of interest. Here we found that the binding affinity of human receptor angiotensin-converting enzyme 2 (hACE2) and receptor-binding domain(RBDs) increased in the order of wild type (Wuhan-strain) < Beta < Alpha < OmicronBA.5 < Gamma < Delta < Omicron BA.2.75 < BA.1 < BA.3 < BA.2. Interactions between docked complexes revealed that the RBD residue positions like 452, 478, 493, 498, 501, and 505 are crucial in creating strong interactions with hACE2. Omicron BA.2 shows the highest binding capacity to the hACE2 receptor among all the mutant complexes. The BA.5's L452R, F486V, and T478K mutation significantly impact the interaction network in the BA.5 RBD-hACE2 interface. Here for the first time, we report the His505, an active residue on the RBD forming a salt bridge in the BA.2, leading to increased mutation stability. When the active RBD residues are mutated, binding affinity and intermolecular interactions increase across all mutant complexes. By examining the differences in different variants, this study may provide a solid foundation for structure-based drug design for newly emerging variants.

Unveiling the Influence of Carbon Nanotube Diameter and Surface Modification on the Anchorage of L-Asparaginase

L-asparaginase (ASNase, EC 3.5.1.1) is an amidohydrolase enzyme known for its anti-cancer properties, with an ever-increasing commercial value. Immobilization has been studied to improve the enzyme’s efficiency, enabling its recovery and reuse, enhancing its stability and half-life time. In this work, the effect of pH, contact time and enzyme concentration during the ASNase physical adsorption onto pristine and functionalized multi-walled carbon nanotubes (MWCNTs and f-MWCNTs, respectively) with different size diameters was investigated by maximizing ASNase relative recovered activity (RRA) and immobilization yield (IY). Immobilized ASNase reusability and kinetic parameters were also evaluated. The ASNase immobilization onto f-MWCNTs offered higher loading capacities, enhanced reusability, and improved enzyme affinity to the substrate, attaining RRA and IY of 100 and 99%, respectively, at the best immobilization conditions (0.4 mg/mL of ASNase, pH 8, 30 min of contact time). In addition, MWCNTs diameter proved to play a critical role in determining the enzyme binding affinity, as evidenced by the best results attained with f-MWCNTs with diameters of 10–20 nm and 20–40 nm. This study provided essential information on the impact of MWCNTs diameter and their surface functionalization on ASNase efficiency, which may be helpful for the development of innovative biomedical devices or food pre-treatment solutions.

Molecular Basis of Mink ACE2 Binding to SARS-CoV-2 and Its Mink-Derived Variants.

ABSTRACTSevere acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is transmitted between humans and minks, and some mutations in the spike (S) protein, especially in the receptor-binding domain (RBD), have been identified in mink-derived viruses. Here, we examined binding of the mink angiotensin-converting enzyme 2 (ACE2) receptor to mink-derived and important human-originating variants, and we demonstrated that most of the RBD variants increased the binding affinities to mink ACE2 (mkACE2). Cryo-electron microscopy structures of the mkACE2-RBD Y453F (with a Y-to-F change at position 453) and mkACE2-RBD F486L complexes helped identify the key residues that facilitate changes in mkACE2 binding affinity. Additionally, the data indicated that the Y453F and F486L mutations reduced the binding affinities to some human monoclonal antibodies, and human vaccinated sera efficiently prevented infection of human cells by pseudoviruses expressing Y453F, F486L, or N501T RBD. Our findings provide an important molecular mechanism for the rapid adaptation of SARS-CoV-2 in minks and highlight the potential influence of the main mink-originating variants for humans.IMPORTANCE Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has a broad range of hosts. Mink-derived SARS-CoV-2 can transmit back to humans. There is an urgent need to understand the binding mechanism of mink-derived SARS-CoV-2 variants to mink receptor. In this study, we identified all mutations in the receptor-binding domain (RBD) of spike (S) protein from mink-derived SARS-CoV-2, and we demonstrated the enhanced binding affinity of mink angiotensin-converting enzyme 2 (ACE2) to most of the mink-derived RBD variants as well as important human-originating RBD variants. Cryo-electron microscopy structures revealed that the Y453F and F486L mutations enhanced the binding forces in the interaction interface. In addition, Y453F and F486L mutations reduced the binding affinities to some human monoclonal antibodies, and the SARS-CoV-2 pseudoviruses with Y453F, F486L, or N501T mutations were neutralized by human vaccinated sera. Therefore, our results provide valuable information for understanding the cross-species transmission mechanism of SARS-CoV-2.

Computational Analysis Reveals that Dual Point Mutation in Rice SBEIIb Leads to Decrease in Starch Binding Affinity

Aim: To check the effect of mutation on the binding affinity of Starch branching enzyme II with maltopentaose substrate.&#x0D; Place and Duration of Study: Department of Bioinformatics, Centre for Plant Molecular Biology and Bioinformatics, Tamil Nadu Agricultural University, Coimbatore, between May 2022 and July 2022.&#x0D; Methodology: The wild type rice SBEIIb and mutated rice SBEIIb protein were modelled using SWISS MODEL and subjected to PROCHECK for validation of the modelled protein. These protein structures were docked with ligand maltopentaose using Auto dock vina of PyRx software. The docked structures were visualized in BIOVIA Discovery studio.&#x0D; Results: There was a decrease in binding affinity after mutating the protein at 2 of the maltopentaose binding sites (Y178A &amp; F234A). The binding energy of wild type rice SBEIIb protein was -7.5kcal mol-1 whereas after mutation it decreased to -5.8 kcal mol-1. The number of hydrogen bonds also decreased from 8 to 4 respectively.&#x0D; Conclusion: Double mutation of binding site residues resulted in binding affinity as well as interactions. In silico analysis prior to wet lab experiments lead to rational choice of mutations that may lead to the production of rice grains with reduced amylopectin content.

Escherichia coli expression and characterization of α-amylase from Geobacillus thermodenitrificans DSM-465.

Alpha amylase, catalyzing the hydrolysis of starch is a ubiquitous enzyme with tremendous industrial applications. A 1698 bp gene coding for 565 amino acid amylase was PCR amplified from Geobacillus thermodenitrificans DSM-465, cloned in pET21a (+) plasmid, expressed in BL21 (DE3) strain of E. coli and characterized. The recombinant enzyme exhibited molecular weight of 63 kDa, optimum pH 8, optimum temperature 70°C, and KM value of 157.7µM. On pilot scale, the purified enzyme efficiently removed up to 95% starch from the cotton fabric indicating its desizing ability at high temperature. 3D model of enzyme built by Raptor-X and validated by Ramachandran plot appeared as a monomer having 31% α-helices, 15% β-sheets, and 52% loops. Docking studies have shown the best binding affinity of enzyme with amylopectin (∆G -10.59). According to our results, Asp 232, Glu274, Arg448, Glu385, Asp34, Asn276, and Arg175 constitute the potential active site of enzyme.

Analysis of ligand binding and resulting conformational changes in pyrophosphatase NUDT9.

Nudix hydrolase 9 (NUDT9) is a member of the nucleoside linked to another moiety X (NUDIX) protein superfamily, which hydrolyses a broad spectrum of organic pyrophosphates from metabolic processes. ADP‐ribose (ADPR) has been the only known endogenous substrate accepted by NUDT9 so far. The Ca2+‐permeable transient receptor potential melastatin subfamily 2 (TRPM2) channel contains a homologous NUDT9‐homology (NUDT9H) domain and is activated by ADPR. Sustained Ca2+ influx via ADPR‐activated TRPM2 triggers apoptotic mechanisms. Thus, a precise regulation of cellular ADPR levels by NUDT9 is essential. A detailed characterization of the enzyme‐substrate interaction would help to understand the high substrate specificity of NUDT9. Here, we analysed ligand binding to NUDT9 using a variety of biophysical techniques. We identified 2′‐deoxy‐ADPR as an additional substrate for NUDT9. Similar enzyme kinetics and binding affinities were determined for the two ligands. The high‐affinity binding was preserved in NUDT9 containing the mutated NUDIX box derived from the human NUDT9H domain. NMR spectroscopy indicated that ADPR and 2′‐deoxy‐ADPR bind to the same binding site of NUDT9. Backbone resonance assignment and subsequent molecular docking allowed further characterization of the binding pocket. Substantial conformational changes of NUDT9 upon ligand binding were observed which might allow for the development of NUDT9‐based ADPR fluorescence resonance energy transfer sensors that may help with the analysis of ADPR signalling processes in cells in the future.

Virtual Screening Based Discovery of PTP1B Inhibitors and Their Biological Evaluations

Background : The discovery of novel antidiabetics for the treatment of type 2 diabetes mellitus (T2DM) is an important task nowadays because the current treatment approaches have certain limitations. The reported studies showed that the protein tyrosine phosphatase 1B (PTP1B) is a valuable target, can be used to develop significant antidiabetic molecules. Objective: In the present investigation, computational methods and biological evaluation studies have been applied to develop novel PTP1B inhibitors with good enzyme binding affinity and activity. Methods: Virtual screening (docking) analysis of SPECS database compounds on PTP1B enzyme was performed using Schrodinger software. In vitro and in vivo biological evaluations had been conducted with the identified hits. Results: The results revealed that the molecules identified through these studies have shown significant interactions with the active site residues of the PTP1B enzyme. The compounds S1 and S2 provided significant binding interactions with the residues (Arg221 and Gln262) and have shown considerable in vitro PTP1B inhibitory activity and in vivo antidiabetic activity. The compounds S1 and S2 possessed 35.44±0.12% and 33.68±0.08% inhibitory activities, respectively. Conclusion: These identified hits will be used as a template for design and development of novel PTP1B inhibitors with a compatible pharmacokinetic profile.